Inhibitors of protein kinases

ABSTRACT

The present invention is directed to compounds of formula I-II and pharmaceutically acceptable salts, esters, and prodrugs thereof which are inhibitors of syk and/or JAK kinase. The present invention is also directed to intermediates used in making such compounds, the preparation of such a compound, pharmaceutical compositions containing such a compound, methods of inhibition syk and/or JAK kinase activity, methods of inhibition the platelet aggregation, and methods to prevent or treat a number of conditions mediated at least in part by syk and/or JAK kinase activity, such as undesired thrombosis and Non Hodgkin&#39;s Lymphoma.

CROSS REFERENCES TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.12/386,848 filed Apr. 22, 2009, which claims the benefit of U.S.Provisional Patent Application No. 61/047,077, filed Apr. 22, 2008; theentire disclosure of each are incorporated herein by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

This invention is directed to pyrimidine, pyrrolopyrimidine andpurine-based analogs which act as inhibitors of Spleen tyrosine kinase(syk) and/or JAK kinases. This invention is also directed topharmaceutical compositions containing the pyrimidine compounds andmethods of using the compounds or compositions to treat a conditioncharacterized by undesired thrombosis. The invention is also directed tomethods of making the compounds described herein.

2. State of the Art

Protein kinases constitute a large family of structurally relatedenzymes that are responsible for the control of a variety of signaltransduction processes within cells (see, e.g., Hardie and Hanks, TheProtein Kinase Facts Book, I and II, Academic Press, San Diego, Calif.,1995). Protein kinases are thought to have evolved from a commonancestral gene due to the conservation of their structure and catalyticfunction. Almost all kinases contain a similar 250-300 amino acidcatalytic domain. The kinases can be categorized into families by thesubstrates they phosphorylate (e.g., protein-tyrosine,protein-serine/threonine, lipids, etc.). Sequence motifs have beenidentified that generally correspond to each of these families (see,e.g., Hanks & Hunter, (1995), FASEB J. 9:576-596; Knighton et al.,(1991), Science 253:407-414; Hiles et al., (1992), Cell 70:419-429; Kunzet al., (1993), Cell 73:585-596; Garcia-Bustos et al., (1994), EMBO J.13:2352-2361).

Many diseases are associated with abnormal cellular responses triggeredby protein kinase-mediated events. These diseases include autoimmunediseases, inflammatory diseases, bone diseases, metabolic diseases,neurological and neurodegenerative diseases, cancer, cardiovasculardiseases, allergies, asthma, alzheimer's disease and hormone-relateddiseases. As a consequence, there has been substantial efforts inmedicinal chemistry to find inhibitors of protein kinases for use astherapeutic agents.

Immunoreceptor tyrosine activation motif (ITAM)-mediated signaling hasemerged as a primary event in signaling pathways responsible for humanpathologies. ITAM-mediated signaling is responsible for relayingactivation signals initiated at classical immune receptors such asT-cell receptors, B-cell receptors, Fc receptors in immune cells and atGPVI and FcγRIIa in platelets to downstream intracellular molecules suchas syk and ZAP-70 (Underhill, D. M and Goodridge, H. S., TrendsImmunol., 28:66-73, 2007).

The binding of a ligand to an ITAM-containing receptor triggerssignaling events which allows for the recruitment of proteins from afamily of nonreceptor tyrosine kinases called the Src family. Thesekinases phosphorylate tyrosine residues within the ITAM sequence, aregion with which the tandem SH2 domains on either syk or ZAP-70interact.

Syk, along with Zap-70, is a member of the syk family of proteintyrosine kinases. The interaction of syk or ZAP-70 with diphosphorylatedITAM sequences induces a conformation change in the kinases that allowsfor tyrosine phosphorylation of the kinase itself. Phosphorylated Sykfamily members activate a multitude of downstream signaling pathwayproteins which include Src homology 2 (SH2) domain containingleukocyte-specific phosphoprotein of 76 kDa (SLP-76), Linker ofActivation of T-cells (LAT) and PLC (phospholipase C)γ2.

Human pathologies attributed to dysfunctional ITAM-mediated signalinginclude autoimmune diseases such as rheumatoid arthritis, systemiclupus, multiple sclerosis, hemolytic anemia, immune-thrombocytopeniapurpura, and heparin-induced thrombocytopenia and arteriosclerosis.Interestingly, many of the above mentioned diseases are thought to occurthrough crosslinking of Fc receptors by antibodies which, via syk,activate a signaling cascade in mast, basophil and other immune cellsthat result in the release of cell mediators responsible forinflammatory reactions. The release of mediators and the production ofcytokines in IgE stimulation-dependent allergic and inflammatoryreactions from mast cells and basophiles can be controlled by inhibitingthe tyrosine kinase activity of syk (Rossi, A. B. et al., J Allergy ClinImmunol., 118:749-755, 2006). In immune-thrombocytopenia, antibody boundplatelets are cleared by the spleen by an Fc receptor/ITAM/syk-mediatedprocess (Crow, A. R. et al., Blood, 106:abstract 2165, 2005).Drug-induced thrombocytopenia, caused by heparin-platelet factor 4immune complexes that activate platelet FcγRIIa, also involve syksignaling downstream of receptor engagement (Reilly, M. P., Blood,98:2442-2447, 2001).

Platelet agonists induce inside-out integrin signaling resulting infibrinogen binding and platelet aggregation. This initiates outside-insignaling which produces further stimulation of platelets. syk isactivated during both phases of integrin signaling, and inhibition ofsyk is shown to inhibit platelet adhesion to immobilized proteins (Law,D. A. et al., Blood, 93:2645-2652, 1999). Release of arachidonic acidand serotonin and platelet aggregation induced by collagen are markedlyinhibited in platelets derived from syk deficient mouse (Poole, A. etal., EMBO J., 16:2333-2341, 1997). Thus syk inhibitors may also possessanticoagulation action.

Because of the role syk plays in Ig-induced platelet activations, it islikely to be important in arteriosclerosis and restenosis.Arteriosclerosis is a class of diseases characterized by the thickeningand hardening of the arterial walls of blood vessels. Although all bloodvessels are susceptible to this serious degenerative condition, theaorta and the coronary arteries serving the heart are most oftenaffected. Arteriosclerosis is of profound clinical importance since itcan increase the risk of heart attacks, myocardial infarctions, strokes,and aneurysms.

The traditional treatment for arteriosclerosis includes vascularrecanalization procedures for less-serious blockages and coronary bypasssurgery for major blockages. A serious shortcoming of intravascularprocedures is that, in a significant number of treated individuals, someor all of the treated vessels restenose (i.e., re-narrow). For example,restenosis of an atherosclerotic coronary artery after PTCA occurs in10-50% of patients undergoing this procedure and subsequently requireseither further angioplasty or a coronary artery bypass graft.Furthermore, restenosis of an atherosclerotic coronary artery afterstenting occurs in 10-20% of patients undergoing this procedure andsubsequently requires repeat treatments to maintain adequate blood flowthrough the affected artery. Restenosis generally occurs in a relativelybrief time period, e.g., roughly less than six months, after treatment.

While the exact hormonal and cellular processes promoting restenosishave not been determined, restenosis is thought to be due in part tomechanical injury to the walls of the blood vessels caused by theballoon catheter or other intravascular device. For example, the processof PTCA, in addition to opening the obstructed artery, also injuresresident coronary arterial smooth muscle cells (SMCs). In response tothis injury, adhering platelets, infiltrating macrophages, leukocytes,or the smooth muscle cells themselves release cell-derived growthfactors such as platelet-derived growth factor (PDGF), with subsequentproliferation and migration of medial SMCs through the internal elasticlamina to the area of the vessel intima. Further proliferation andhyperplasia of intimal SMCs and, most significantly, production of largeamounts of extracellular matrix over a period of three to six monthsresults in the filling in and narrowing of the vascular space sufficientto significantly obstruct blood flow.

In addition to the role syk plays in Ig-induced platelet activations,syk plays a very important role in collagen-mediated signaling. Theprimary adhesive protein responsible for platelet adhesion andactivation is collagen. Collagen is a filamentous protein containedwithin the fibrotic caps of atheromas which becomes exposed to bloodduring plaque rupture. Collagen functions initially by binding vonWillebrand factor which tethers platelets through binding plateletmembrane GPIb. Collagen functions secondarily by engaging the twocollagen receptors on platelets, GPVI and integrin α2β1.

GPVI exists in platelet membranes as a complex with FcRγ, an interactionrequired for the expression of GPVI. Activation of FcγRIIa on plateletsresults in platelet shape change, secretion and thrombosis. Signaling bythe GPVI/FcRγcomplex is initiated by tyrosine phosphorylation of theITAM domain of FCRγ followed by the recruitment of syk. Activation ofGPVI leads to induction of multiple platelet functions including:activation of integrins α2β1 to achieve firm platelet adhesion, and GPIIb-IIIa which mediates platelet aggregation and thrombosis growth;platelet secretion, allowing for the delivery of inflammatory proteinssuch as CD40L, RANTES and TGFβ to the vessel wall; and the expression ofP-selectin which allows for the recruitment of leukocytes. Therefore, itis believed that syk inhibitors can inhibit thrombotic events mediatedby platelet adhesion, activation and aggregation.

It has been reported that the tyrosine phosphorylation of intracellularprotein (activation) induced by stimulation of a receptor for IgGantibody, FcγR, and the phagocytosis mediated by FcγR are considerablyinhibited in macrophages derived from syk deficient mouse (Crowley, M.T. et al., J. Exp. Med., 186:1027-1039, 1997). This suggests that sykhas a markedly important role in the FcγR-mediated phagocytosis ofmacrophages.

It has also been reported that an antisense oligonucleotide of syksuppresses the apoptosis inhibition of eosinophils induced by GM-CSF(Yousefi, S. et al., J. E. Med., 183:1407-1414, 1996), showing that sykis essential for the life extending signal of eosinophils caused byGM-CSF and the like. Since life extension of eosinophils is closelyrelated to the transition of diseases into a chronic state in allergicdisorders, such as asthma, syk inhibitors can also serve as therapeuticagents for chronic eosinophilic inflammation.

Syk is important for the activation of B-cells via a B-cell antigenreceptor and is involved in the phosphatidylinositol metabolism andincrease in the intracellular calcium concentration caused by theantigen receptor stimulation (Hutchcroft, J E. et al., J. Biol. Chem.,267:8613-8619, 1992; and Takata, M. et al., EMBO J., 13:1341-1349,1994). Thus, syk inhibitors may be used to control the function ofB-cells and are, therefore, expected to serve as therapeutic agents forantibody-related diseases.

Syk binds to a T-cell antigen receptor, quickly undergoes tyrosinephosphorylation through crosslinking of the receptor and synergisticallyacts upon intracellular signals mediated by Src tyrosine kinases such asLck (Couture, C. et al., Proc. Natl. Acad. Sci. USA, 91:5301-5305, 1994;and Couture, C. et al., Mol. Cell. Biol., 14:5249-5258, 1994). syk ispresent in mature T-cell populations, such as intraepithelial γδ T-cellsand naïve αβ T-cells, and has been reported to be capable ofphosphorylation of multiple components of the TCR signaling cascade(Latour, S. et. al., Mol Cell Biol., 17:4434-4441, 1997). As aconsequence, syk inhibitors may serve as agents for inhibiting cellularimmunity mediated by T-cell antigen receptor.

Recent comparative genomic hybridization studies have identified syk asanother gene important in the pathogenesis of Mantle Cell Lymphoma (MCL)(Chen, R. et al. Journal of Clinical Oncology, 2007 ASCO Annual MeetingProceedings (Post-Meeting Edition). Vol 25, No 18S (June 20 Supplement),2007: 8056). MCL represents 5-10% of all non-Hodgkins lymphomas and itis a difficult form of lymphoma to treat. It has the worst prognosisamong the B cell lymphomas with median survival of three years. It hasbeen reported that Syk is overexpressed in MCL (Rinaldi, A, et. al, Br.J. Haematol., 2006; 132:303-316) and that Syk mediates mTOR (mammaliantarget of Rapamycin) survival signals in follicular, mantel cell,Burkitt's, and diffuse large B-cell non-Hodgkin's lymphomas (Leseux, L.,et. al, Blood, 2006; 108:4156-4162).

Several lines of evidence suggest that many B-cell lymphomas depend uponB-cell receptor (BCR)-mediated survival signals. BCR signaling inducesreceptor oligomerization and phosphorylation of Igα and β immunoreceptortyrosine-based activated motifs by SRC family kinases. ITAMphosphorylation results in the recruitment and activation of syk thatinitiates downstream events and amplifies the original BCR signal. Giventhe role of tonic BCR signaling in normal B cell and syk-dependentsurvival of non-Hodgkins lymphoma cell lines in vitro (Chen, L., et. al,Blood, 2006; 108:3428-3433), syk inhibition is a promising rationaltreatment target for certain B-cell lymphomas and chronic lymphocyticleukemia (CLL) (Stefania Gobessi, Luca Laurenti, Pablo Longo, LauraCarsetti, Giuseppe Leone, Dimitar G. Efremov, Constitutive activation ofthe protein tyrosine kinase Syk in Chronic Lymphocytic Leukemia B-cells,Blood, 2007, 110, Abstract 1123). Recent data shows that administrationof a multikinase inhibitor which inhibits syk, may have significantclinical activity in CLL patients (Friedberg J W et al, Blood 2008;112(11), Abstract 3).

The oncogenic potential of the spleen tyrosine kinase (Syk) has beendescribed in a number of different settings. Clinically, Sykover-expression is reported in Mantle Cell Lymphoma (Rinaldi, A, et. al,Br. J. Haematol., 2006; 132:303-316) and the TEL-Syk fusion protein(Translocated ETS Leukemia) generated by a chromosomal translocation(t(9;12)(q22;p12)) leads to increased Syk activity and is associatedwith myelodysplastic syndrome (Kuno, Y., et. al, Blood, 2001;97:1050-1055). Leukemia is induced in mice by adoptively transferringbone marrow cells that express human TEL-Syk (Wossning, T., JEM, 2006;203:2829-2840). Further, in mouse primary bone marrow cells,over-expression of Syk results in IL-7 independent growth in culture(Wossning, T., et. al, JEM, 2006; 203:2829-2840).

Interestingly, Syk signaling appears to be required for B-celldevelopment and survival in humans and mouse. Inducible loss of theB-cell receptor (Lam, K., et. al, Cell, 1997; 90:1073-1083) or Igα(Kraus, M., et. al, Cell, 2004; 117:787-800) results in loss ofperipheral B-cells in mice. Over-expression of the protein tyrosinephosphatase PTP-RO, which is known to negatively regulate Syk activity,inhibits proliferation and induces apoptosis in cell lines derived fromnon-Hodgkin's lymphomas (Chen, L., et. al, Blood, 2006; 108:3428-3433).Finally, B-cell lymphomas rarely exhibit loss of BCR expression, andanti-idiotype therapy rarely leads to resistance (Kuppers, R. Nat RevCancer, 2005; 5:251-262).

Engagement of the antigen-specific B cell receptor (BCR) activatesmultiple signaling pathways that ultimately regulate the cellsactivation status, promoting survival and clonal expansion. Signalingthrough the BCR is made possible by its association with two othermembers of the immunoglobulin super-family; Igα and Igβ, each bearing animmuno-tyrosine based activation motif (ITAM) (Jumaa, Hendriks et al.Annu Rev Immunol 23: 415-45 (2005). The ITAM domain is directlyphosphorylated by Src family kinases in response to BCR engagement. Thespleen tyrosine kinase (Syk) docks with and phosphorylates the ITAM, aprocess that enhances its kinase activity, resulting in Sykautophosphorylation and tyrosine phosphorylation of multiple downstreamsubstrates (Rolli, Gallwitz et al. Mol Cell 10(5): 1057-69 (2002). Thissignaling pathway is active in B cells beginning at the transition frompro- to pre-B cell stage of development, when the newly formed pre-BCRis expressed. In fact, B cell development arrests at the pro-B cellstage in Syk knockout mice (Cheng, Rowley et al. 1995; Turner, Mee etal. Nature 378(6554): 303-6 (1995). Inducible loss of the B cellreceptor (Lam, Kuhn et al. Cell 90(6): 1073-83 (1997) or Igα (Kraus,Alimzhanov et al. Cell 117(6): 787-800 (2004) results in loss ofperipheral B cells in mice. Human B cells also appear to require Syk forproliferation and survival. Over-expression of the protein tyrosinephosphatase PTP-RO, a negative regulator of Syk activity, inhibitsproliferation and induces apoptosis in cell lines derived fromnon-Hodgkin's lymphomas (NHL) (Chen, Juszczynski et al. Blood 108(10):3428-33 (2006). Knock down of Syk by siRNA in the NHL line SUDHL-4 ledto a block in the G1/S transition of the cell cycle (Gururajan, Dasu etal. J Immunol 178(1): 111-21 (2007). Together, these data suggest thatSyk signaling is required for the development, proliferation, and evensurvival of human and mouse B cells.

Conversely, the oncogenic potential of Syk has been described in anumber of different settings. Clinically, Syk over-expression isreported in Mantle Cell Lymphoma (Rinaldi, Kwee et al. Br J Haematol132(3): 303-16 (2006) and the TEL-Syk fusion protein (Translocated ETSLeukemia) generated by a chromosomal translocation (t(9;12)(q22;p12))leads to increased Syk activity and is associated with myelodysplasticsyndrome (Kuno, Abe et al. Blood 97(4): 1050-5 (2001). Leukemia isinduced in mice by the adoptive transfer of bone marrow cells thatexpress human TEL-Syk (Wossning, Herzog et al. J Exp Med 203(13):2829-40 (2006). Further, in mouse primary bone marrow cells,over-expression of Syk results in IL-7 independent growth in culture(Wossning, Herzog et al. 2006). Consistently, Syk was reported tomediate mTOR (mammalian target of Rapamycin) survival signals infollicular, mantle cell, Burkitt's, and diffuse large B-cell NHL(Leseux, Hamdi et al. Blood 108(13): 4156-62 (2006). Additional recentstudies also suggest that Syk-dependant survival signals may play a rolein B-cell malignancies, including DLBCL, mantle cell lymphoma andfollicular lymphoma (Gururajan, Jennings et al. 2006; Irish, Czerwinskiet al. J Immunol 176(10): 5715-9 (2006). Given the role of tonic BCRsignaling in normal B cells and Syk-dependent survival of NHL cell linesin vitro, the specific inhibition of Syk may prove promising for thetreatment of certain B-cell lymphomas.

Recently, R406 (Rigel Pharmaceuticals) was reported to inhibit ITAMsignaling in response to various stimuli, including FcεR1 and BCRinduced Syk activation (Braselmann, Taylor et al. J Pharmacol Exp Ther319(3): 998-1008 (2006). Interestingly, this ATP-competitive inhibitorof Syk was also active against Flt3, cKit, and JAK kinases, but notagainst Src kinsase (Braselmann, Taylor et al. 2006). Activatingmutations to Flt3 are associated with AML and inhibition of this kinaseis currently under clinical development (Burnett and Knapper HematologyAm Soc Hematol Educ Program 2007: 429-34 (2007). Over-activation of thetyrosine kinase cKit is also associated with hematologic malignancies,and a target for cancer therapy (Heinrich, Griffith et al. Blood 96(3):925-32 (2000). Similarly, JAK3 signaling is implicated in leukemias andlymphomas, and is currently exploited as a potential therapeutic target(Heinrich, Griffith et al. 2000). Importantly, the multi-kinaseinhibitory activity of R406 attenuates BCR signaling in lymphoma celllines and primary human lymphoma samples, resulting in apoptosis of theformer (Chen, Monti et al. Blood 111(4): 2230-7 (2008). Further, a phaseII clinical trial reported favorable results by this compound inrefractory NHL and chronic lymphocytic leukemia (Friedberg J W et al,Blood 2008; 112(11), Abstract 3). Although the precise mechanism ofaction is unclear for R406, the data suggest that inhibition of kinasesthat mediate survival signaling in lymphocytes is clinically beneficial.

Additional recent studies also suggest that syk-dependant survivalsignals may play a role in B-cell malignancies, including DLBCL, mantlecell lymphoma and follicular lymphoma (see e.g., S. Linfengshen et al.Blood, February 2008; 111: 2230-2237; J. M. Irish et al. Blood, 2006;108: 3135-3142; A. Renaldi et al. Brit J. Haematology, 2006; 132:303-316; M. Guruoajan et al. J. Immunol, 2006; 176: 5715-5719; L. Laseuxet al. Blood, 2006; 108: 4156-4162.

JAK kinases (Janus Kinases) are a family of cytoplasmic protein tyrosinekinases including JAK1, JAK2, JAK3 and TYK2. The JAKs play a crucialrole in cytokine signaling. Each of the JAK kinases is selective for thereceptors of certain cytokines, though multiple JAK kinases can beaffected by particular cytokine or signaling pathways. Studies suggestthat JAK3 associates with the common cytokine receptor gamma chain (Fcγor γc) of the various cytokine receptors. JAK3 in particular selectivelybinds to receptors and is part of the cytokine signaling pathway for andactivated by IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21. JAK1 interactswith, among others, the receptors for cytokines IL-2, IL-4, IL-7, IL-9and IL-21, while JAK2 interacts with, among others, the receptors forIL-9 and TNF-α. Upon the binding of certain cytokines to their receptors(e.g., IL-2, IL-4, IL-7, IL-9, IL-15 and IL-21), receptoroligomerization occurs, resulting in the cytoplasmic tails of associatedJAK kinases being brought into proximity and facilitating thetrans-phosphorylation of tyrosine residues on the JAK kinase. Thistrans-phosphorylation results in the activation of the JAK kinase.

The downstream substrates of JAK family kinases include the signaltranducer activator of transcription (STAT) proteins. Phosphorylated JAKkinases bind various STAT (Signal Transducer and Activator ofTranscription) proteins. STAT proteins, which are DNA binding proteinsactivated by phosphorylation of tyrosine residues, function both assignaling molecules and transcription factors and ultimately bind tospecific DNA sequences present in the promoters of cytokine-responsivegenes (Leonard et al., (2000), J. Allergy Clin. Immunol. 105:877-888).

JAK/STAT signaling has been implicated in the mediation of many abnormalimmune responses such as allergies, asthma, autoimmune diseases such astransplant (allograft) rejection, rheumatoid arthritis, amyotrophiclateral sclerosis and multiple sclerosis, as well as in solid andhematologic malignancies such as leukemia and lymphomas. For a review ofthe pharmaceutical intervention of the JAK/STAT pathway see Frank,(1999), Mol. Med. 5:432:456 and Seidel et al., (2000), Oncogene19:2645-2656.

JAK3 in particular has been implicated in a variety of biologicalprocesses. For example, the proliferation and survival of murine mastcells induced by IL-4 and IL-9 have been shown to be dependent on JAK3-and gamma chain-signaling (Suzuki et al., (2000), Blood 96:2172-2180).JAK3 also plays a crucial role in IgE receptor-mediated mast celldegranulation responses (Malaviya et al., (1999), Biochem. Biophys. Res.Commun 257:807-813), and inhibition of JAK3 kinase has been shown toprevent type I hypersensitivity reactions, including anaphylaxis(Malaviya et al., (1999), J. Biol. Chem. 274:27028-27038). JAK3inhibition has also been shown to result in immune suppression forallograft rejection (Kirken, (2001), Transpl. Proc. 33:3268-3270). JAK3kinases have also been implicated in the mechanism involved in early andlate stages of rheumatoid arthritis (Muller-Ladner et al., (2000), J.Immunal. 164:3894-3901); familial amyotrophic lateral sclerosis (Trieuet al., (2000), Biochem Biophys. Res. Commun. 267:22-25); leukemia(Sudbeck et al., (1999), Clin. Cancer Res. 5:1569-1582); mycosisfungoides, a form of T-cell lymphoma (Nielsen et al., (1997), Prac.Natl. Acad. Sci. USA 94:6764-6769); and abnormal cell growth (Yu et al.,(1997), J. Immunol. 159:5206-5210; Catlett-Falcone et al., (1999),Immunity 10:105-115).

JAK1, JAK2, and TYK2 are expressed ubiquitously, whereas JAK3 isexpressed predominantly in hematopoietic cells. The JAK kinases,including JAK3, are abundantly expressed in primary leukemic cells fromchildren with acute lymphoblastic leukemia, the most common form ofchildhood cancer, and studies have correlated STAT activation in certaincells with signals regulating apoptosis (Demoulin et al., (1996), Mol.Cell. Biol. 16:4710-6; Jurlander et al., (1997), Blood. 89:4146-52;Kaneko et al., (1997), Clin. Exp. Immun 109:185-193; and Nakamura etal., (1996), J. Biol. Chem. 271: 19483-8). They are also known to beimportant for lymphocyte differentiation, function and survival. JAK-3in particular plays an essential role in the function of lymphocytes,macrophages, and mast cells. Given the importance of this JAK kinase,compounds which modulate the JAK pathway, including those selective forJAK3, can be useful for treating diseases or conditions where thefunction of lymphocytes, macrophages, or mast cells is involved (Kudlaczet al., (2004) Am. J. Transplant 4:51-57; Changelian (2003) Science302:875-878). Conditions in which targeting of the JAK pathway ormodulation of the JAK kinases, particularly JAK3, are contemplated to betherapeutically useful include, leukemia, lymphoma, transplant rejection(e.g., pancreas islet transplant rejection, bone marrow transplantapplications (e.g., graft-versus-host disease), autoimmune diseases(e.g., diabetes, rheumatoid arthritis, lupus, psoriasis), andinflammation (e.g., asthma, allergic reactions). Conditions which canbenefit from JAK3 inhibition are discussed in greater detail below.Recent data on JAK inhibition has been reported in kidney allograftpatients treated with CP-690,550 and showed that markers of allogeneicresponse (interferon gamma) can be reduced (Van Gurp E A et al (2009)Transplanatation 87:79-86).

In view of the numerous conditions that are contemplated to benefit bytreatment involving modulation of the JAK pathway it is immediatelyapparent that new compounds that modulate JAK pathways and methods ofusing these compounds should provide substantial therapeutic benefits toa wide variety of patients. Provided herein are novel2,4-pyrimidinediamine, pyrrolopyrimidine and purine-based compounds foruse in the treatment of conditions in which targeting of the JAK pathwayor inhibition of JAK kinases, particularly JAK3, are therapeuticallyuseful.

Patents and patent applications related to modulation of the JAK pathwayinclude: U.S. Pat. Nos. 5,728,536; 6,080,747; 6,080,748; 6,133,305;6,177,433; 6,210,654; 6,313,130; 6,316,635; 6,433,018; 6,486,185;6,506,763; 6,528,509; 6,593,357; 6,608,048; 6,610,688; 6,635,651;6,677,368; 6,683,082; 6,696,448; 6,699,865; 6,777,417; 6,784,195;6,825,190; 6,506,763; 6,784,195; 6,528,509; 6,608,048; 7,105,529;6,699,865; 6,825,190; 6,815,439; 6,949,580; 7,056,944; 6,998,391;7,074,793; 6,969,760; U.S. Pat. App. Pub. No. 2001/0007033 A1;2002/0115173 A1; 2002/0137141 A1; 2003/0236244 A1; 2004/0102455 A1;2004/0142404 A1; 2004/0147507 A1; and 2004/0214817 A1; and Internationalpatent applications WO 95/03701A1; WO 99/15500A1; WO 00/00202A1; WO00/10981A1; WO 00/47583A1; WO 00/51587A2; WO 00/55159A2; WO 01/42246A2;WO 01/45641A2; WO 01/52892A2; WO 01/56993A2; WO 01/57022A2; WO01/72758A1; WO 02/00661A1; WO 02/43735A1; WO 02/48336A2; WO 02/060492A1;WO 02/060927A1; WO 02/096909A1; WO 02/102800A1; WO 03/020698A2; WO03/048162A1; WO 03/101989A1; WO 2004/016597A2; WO 2004/041789A1; WO2004/041810A1; WO 2004/041814A1; WO 2004/046112A2; WO 2004/046120A2; WO2004/047843A1; WO 2004/058749A1; WO 2004/058753A1; WO 2004/085388A2; WO2004/092154A1; WO 2005/009957A1; WO 2005/016344A1; WO 2005/028475A2; andWO 2005/033107A1.

Patents and patent applications describing substituted pyrimidinediaminecompounds include: U.S. application Ser. No. 10/355,543 filed Jan. 31,2003 (US2004/0029902A1), international application Serial No.PCT/US03/03022 filed Jan. 31, 2003 (WO 03/063794), U.S. application Ser.No. 10/631,029 filed Jul. 29, 2003, international application Serial No.PCT/US03/24087 (WO 04/014382), U.S. application Ser. No. 10/903,263filed Jul. 30, 2004, and international application Serial No.PCT/US2004/24716 (WO 05/016893), the disclosures of which areincorporated herein by reference. Substituted pyrimidinediaminecompounds are also described in international patent applicationpublication numbers: WO 02/059110, WO 03/074515, WO 03/106416, WO03/066601, WO 03/063794, WO 04/046118, WO 05/016894, WO 05/122294, WO05/066156, WO 03/002542, WO 03/030909, WO 00/39101, WO 05/037800 andU.S. Pat. Pub. No. 2003/0149064.

While progress has been made in this field, there remains a need in theart for compounds that inhibit syk and/or JAK kinase, as well as formethods for treating conditions in a patient, such as restenosis,thrombosis, and/or inflammation that can benefit from such inhibition.Moreover, the availability of compounds that selectively inhibit one ofthese kinases as compared to other kinases would also be desirable. Thepresent invention satisfies this and other needs.

BRIEF SUMMARY OF THE INVENTION

The present invention provides novel compounds having activity asinhibitors of syk activity (also referred to herein as “syk inhibitors”)and/or JAK kinase activity (also referred to herein as “JAKinhibitors”), as well as to methods for their preparation and use, andto pharmaceutical compositions containing the same. Such compounds havethe following structure (I-II):

or a pharmaceutically acceptable salt thereof, wherein Y^(1a), Z^(1a),R^(1a), R^(1c), R^(2a), R^(2c), R^(3a), R^(3c), R^(4a), R^(4b), R^(4d),R^(5a), R^(6a), R^(7a) and R^(7c) are as defined below.

The present invention also provides a pharmaceutical compositioncomprising a therapeutically effective amount of a compound of formulaI-II, or a pharmaceutical acceptable salt thereof, and apharmaceutically acceptable carrier and/or diluent.

The compounds of the present invention have utility over a wide range oftherapeutic applications, and may be used to treat a variety ofconditions, mediated at least in part by syk activity, in both men andwomen, as well as a mammal in general (also referred to herein as a“subject”). For example, such conditions include, but are not limitedto, those associated with cardiovascular disease, inflammatory diseaseor autoimmune disease. More specifically, the compounds of the presentinvention have utility for treating conditions or disorders including,but not limited to: restenosis, thrombosis, inflammation, heparininduced thrombocytopenia, dilated cardiomyopathy, sickle cell disease,atherosclerosis, myocardial infarction, vascular inflammation, unstableangina, acute coronary syndromes, allergy, asthma, rheumatoid arthritis,B-cell mediated diseases such as Non Hodgkin's lymphoma,anti-phospholipid syndrome, lupus, psoriasis, multiple sclerosis, endstage renal disease, hemolytic anemia, immune thrombocytopenic purpura,and chronic lymphocytic leukemia. Thus, in one embodiment, methods aredisclosed which include the administration of an effective amount of acompound of formula (I-II), typically in the form of a pharmaceuticalcomposition, to a subject in need thereof.

The conditions associated with cardiovascular disease is selected fromthe group consisting of acute coronary syndrome, myocardial infarction,unstable angina, refractory angina, occlusive coronary thrombosisoccurring post-thrombolytic therapy or post-coronary angioplasty, athrombotically mediated cerebrovascular syndrome, embolic stroke,thrombotic stroke, transient ischemic attacks, venous thrombosis, deepvenous thrombosis, pulmonary embolism, coagulopathy, disseminatedintravascular coagulation, thrombotic thrombocytopenic purpura,thromboangiitis obliterans, thrombotic disease associated withheparin-induced thrombocytopenia, thrombotic complications associatedwith extracorporeal circulation, thrombotic complications associatedwith instrumentation such as cardiac or other intravascularcatheterization, intra-aortic balloon pump, coronary stent or cardiacvalve, and conditions requiring the fitting of prosthetic devices.

The present invention also provides a method for inhibiting the sykactivity of a blood sample comprising contacting said sample with acompound of the present invention.

The present invention further provides compounds in purified forms, aswell as chemical intermediates.

These and other aspects, objects, features and advantages of theinvention will be apparent upon reference to the following detaileddescription and figures. To this end, various references are set forthherein which describe in more detail certain background information,procedures, compounds and/or compositions, and are each herebyincorporated by reference in their entirety.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows how Syk serves as a key mediator of Fc receptor mediatedsignaling in cellular biology and multiple diseases.

FIG. 2 shows how gene targeting of Syk indicated that Syk serves as akey mediator in arterial platelet biology and a selective target fortreating arterial thrombosis.

FIGS. 3-5 show a general synthesis of compounds of the presentinvention.

FIG. 6 shows a graph of the effect of increasing dose of a compound ofthe present invention on pSTAT6 formation in response to IL4 stimulationof B Ramos cells.

FIG. 7 provides table 1 illustrating compounds of the present inventionand syk IC₅₀s.

FIG. 8 provides table 2 illustrating compounds of the present inventionand SYK IC₅₀s.

FIG. 9 shows specificity data for selected compounds.

FIG. 10 shows the selective inhibition of selected bicyclic compounds.

DETAILED DESCRIPTION OF THE INVENTION

As used herein, the below terms have the following meanings unlessspecified otherwise:

1. ABBREVIATIONS AND DEFINITIONS

The abbreviations used herein are conventional, unless otherwisedefined. The following abbreviations are used: AcOH=acetic acid,AIBN=azobisisobutyronitrile (also azobisisobutylonitrile), aq.=aqueous,Boc=t-butylcarboxy, Bz-benzyl,BOP=benzotriazol-1-yloxytris(dimethylamino)-phosphoniumhexafluorophosphate, BPO=benzoyl peroxide, nBuOH=n-butanol,CBr₄=tetrabromomethane, mCPBA=m-chloroperoxybenzoic acid, CH₂Cl₂ orDCM=dichloromethane, Cs₂CO₃=cesium carbonate, CuCl₂=copper chloride;DIBAL=diisobutylaluminum hydride, DIEA=Hunig's base or diisopropylethylamine, DME=dimethyl ether, DMF=dimethyl formamide, DMSO=dimethylsulfoxide, DPPA=diphenyl phosphoryl azide, Et₃N=triethylamine,EtOAc=ethyl acetate, g=gram, HATU=2-(1H7-Azabenzotriazol-1-yl)-1,1,3,3-tetramethyl uronium hexafluorophosphate,H₂=hydrogen; H₂O=water; HBr=hydrogen bromide; HCl=hydrogen chloride,HIV=human immunodeficiency virus, HPLC=high pressure liquidchromatography, h=hour, IgE=immunoglobulin E, IC₅₀=The concentration ofan inhibitor that is required for 50% inhibition of an enzyme in vitro,IPA=isopropyl alcohol, kg=kilogram, KCN=potassium cyanide, KOH=potassiumhydroxide, K₂PO₄=potassium phosphate, LDA=lithium diisopropylamine,LiAlH₄=lithium aluminum hydride=LiOH: lithium hydroxide;MeCN=acetonitrile; MS=Mass Spec, m/z=mass to charge ratio, MHz=MegaHertz, MeOH=methanol, μM=micromolar, μL=microliter, mg=milligram,mm=millimeter, mM=millimolar, mmol=millimole, mL=milliliter,mOD/min=millioptical density units per minute, min=minute, M=molar,Na₂CO₃=sodium carbonate, ng=nanogram, NaHCO₃=sodium bicarbonate;NaNO₂=sodium nitrite; NaOH=sodium hydroxide; Na₂S₂O₃=sodium bisulfate;Na₂SO₄=sodium sulfate; NBS=N-bromosuccinamide; NH₄Cl=ammonium chloride;NH₄OAc=ammonium acetate; NaSMe=sodium methylthiolate,NBS=N-bromosuccinamide, n-BuLi=n-butyl lithium, nm=nanometer,nM=nanomolar, N=Normal, NMP=N-methylpyrrolidine, NMR=nuclear magneticresonance, Pd/C=palladium on carbon,Pd(PPh₃)₄=Tetrakis-(triphenyl-phosphine)-palladium, pM=picomolar,Pin=pinacolato, PEG=polyethylene glycol, PPh₃ or Ph₃P=triphenylphosphine, RLV=Raucher leukemia virus, Ra—Ni=Rainey Nickel,SOCl₂=thionyl chloride, RT=room temperature, TEA=triethylamine,THF=tetrahydrofuran, TFA=trifluoroacetic acid, TLC=thin layerchromatography, TMS=trimethylsilyl, Tf=trifluoromethylsulfonyl andTSC=trisodium citrate.

It is noted here that as used in this specification and the appendedclaims, the singular forms “a,” “an,” and “the” include plural referenceunless the context clearly dictates otherwise.

“Alkyl,” by itself or as part of another substituent, means, unlessotherwise stated, a straight or branched chain, fully saturatedaliphatic hydrocarbon radical having the number of carbon atomsdesignated. For example, “C₁₋₈alkyl” refers to a hydrocarbon radicalstraight or branched, containing from 1 to 8 carbon atoms that isderived by the removal of one hydrogen atom from a single carbon atom ofa parent alkane. The phrase “unsubstituted alkyl” refers to alkyl groupsthat do not contain groups other than fully saturated aliphatichydrocarbon radicals. Thus the phrase includes straight chain alkylgroups such as methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl,octyl, nonyl, decyl, undecyl, dodecyl and the like. The phrase alsoincludes branched chain isomers of straight chain alkyl groups such asisopropyl, t-butyl, isobutyl, sec-butyl, and the like. Representativealkyl groups include straight and branched chain alkyl groups having 1,2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 carbon atoms. Furtherrepresentative alkyl groups include straight and branched chain alkylgroups having 1, 2, 3, 4, 5, 6, 7 or 8 carbon atoms.

“Alkenyl” by itself or as part of another substituent refers to astraight or branched chain, which may be mono- or polyunsaturated,having the number of carbon atoms designated. For example, “C₂-C₈alkenyl” means an alkenyl radical having from 2, 3, 4, 5, 6, 7 or 8atoms that is derived by the removal of one hydrogen atom from a singlecarbon atom of a parent alkane. Examples include, but are not limited tovinyl, 2-propenyl i.e.—CH═C(H)(CH₃), —CH═C(CH₃)₂, —C(CH₃)═C(H)₂,—C(CH₃)═C(H)(CH₃), —C(CH₂CH₃)═CH₂, butadienyl e.g. 2-(butadienyl),pentadienyl e.g. 2,4-pentadienyl and 3-(1,4-pentadienyl), andhexadienyl, among others, and higher homologs and stereoisomers thereof.A “substituted” alkenyl group includes alkenyl groups in which anon-carbon or non-hydrogen atom is bonded to a carbon double bonded toanother carbon and those in which one of the non-carbon or non-hydrogenatoms is bonded to a carbon not involved in a double bond to anothercarbon. Each site of unsaturation may be either cis or transconfiguration about the double bond(s).

The term “alkynyl”, by itself or as part of another substituent, means astraight or branched chain hydrocarbon radical, which may be mono- orpolyunsaturated, having the number of carbon atoms designated. Forexample, “C₂-C₈ alkynyl” means an alkynyl radical having from 2 to 8carbon atoms that is derived by the removal of one hydrogen atom from asingle carbon atom of a parent alkane. “Unsubstituted alkynyl” refers tostraight and branched chain groups such as those described with respectto unsubstituted alkyl groups as defined above, except that at least onetriple bond exists between two carbon atoms. Examples include, but arenot limited to ethynyl e.g. —C≡C(H), 1-propynyl e.g. —C≡C(CH₃),—C≡C(CH₂CH₃), —C(H₂)C≡C(H), —C(H)₂C≡C(CH₃), and —C(H)₂C≡C(CH₂CH₃) amongothers, and higher homologs and isomers thereof. A “substituted” alkynylgroup includes alkynyl groups in which a non-carbon or non-hydrogen atomis bonded to a carbon triple bonded to another carbon and those in whicha non-carbon or non-hydrogen atom is bonded to a carbon not involved ina triple bond to another carbon.

“Alkylene” by itself or as part of another substituent means a divalentradical derived from an alkane, as exemplified by —CH₂CH₂CH₂CH₂—.Typically, an alkylene group will have from 1, 2, 3, 4, 5, 6, 7 or 8carbon atoms that is derived by the removal of one hydrogen atom from asingle carbon atom of a parent alkyl.

“Cycloalkyl” or “carbocycle”, by themselves or in combination with otherterms, represent, unless otherwise stated, cyclic versions of “alkyl”,“alkenyl” and “alkynyl” in which all ring atoms are carbon. “Cycloalkyl”or “carbocycle” refers to a mono- or polycyclic group. When used inconnection with cycloalkyl substituents, the term “polycyclic” refersherein to fused and non-fused alkyl cyclic structures. “Cycloalkyl” or“carbocycle” may form a bridged ring or a spiro ring. The cycloalkylgroup may have one or more double or triple bond(s). The term“cycloalkenyl” refers to a cycloalkyl group that has at least one siteof alkenyl unsaturation between the ring vertices. The term“cycloalkynyl” refers to a cycloalkyl group that has at least one siteof alkynyl unsaturation between the ring vertices. When “cycloalkyl” isused in combination with “alkyl”, as in C₃₋₈cycloalkylC₃₋₈alkylene-, thecycloalkyl portion is meant to have the stated number of carbon atoms(e.g., from three to eight carbon atoms), while the alkylene portion hasfrom one to eight carbon atoms. Typical cycloalkyl substituents havefrom 3 to 8 ring atoms. Examples of cycloalkyl include cyclopentyl,cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and the like.

“Aryl” by itself or as part of another substituent refers to apolyunsaturated, aromatic, hydrocarbon group containing from 6 to 14carbon atoms, which can be a single ring or multiple rings (up to threerings) which are fused together or linked covalently. Thus the phraseincludes, but is not limited to, groups such as phenyl, biphenyl,anthracenyl, naphthyl by way of example. Non-limiting examples ofunsubstituted aryl groups include phenyl, 1-naphthyl, 2-naphthyl and4-biphenyl. “Substituted aryl group” includes, for example, —CH₂OH (onecarbon atom and one heteroatom replacing a carbon atom) and —CH₂SH. Theterm “heteroalkylene” by itself or as part of another substituent meansa divalent radical derived from heteroalkyl, as exemplified by—CH²⁻CH²⁻S—CH₂CH²⁻— and —CH²⁻S—CH²⁻CH²⁻NH—CH²⁻. For heteroalkylenegroups, heteroatoms can also occupy either or both of the chain termini(e.g., alkyleneoxy, alkylenedioxy, alkyleneamino, alkylenediamino, andthe like). Still further, for alkylene and heteroalkylene linkinggroups, no orientation of the linking group is implied.

The terms “heterocycle”, “heterocyclyl” or “heterocyclic” refer to asaturated or unsaturated non-aromatic cyclic group containing at leastone heteroatom. As used herein, the term “heteroatom” is meant toinclude oxygen (O), nitrogen (N), sulfur (S) and silicon (Si). Eachheterocycle can be attached at any available ring carbon or heteroatom.Each heterocycle may have one or more rings. When multiple rings arepresent, they can be fused together or linked covalently. Eachheterocycle typically contains 1, 2, 3, 4 or 5, independently selectedheteroatoms. Preferably, these groups contain 1, 2, 3, 4, 5, 6, 7, 8, 9or 10 carbon atoms, 0, 1, 2, 3, 4 or 5 nitrogen atoms, 0, 1 or 2 sulfuratoms and 0, 1 or 2 oxygen atoms. More preferably, these groups contain1, 2 or 3 nitrogen atoms, 0-1 sulfur atoms and 0-1 oxygen atoms.Non-limiting examples of heterocycle groups include morpholin-3-one,piperazine-2-one, piperazin-1-oxide, pyridine-2-one, piperidine,morpholine, piperazine, isoxazoline, pyrazoline, imidazoline,pyrazol-5-one, pyrrolidine-2,5-dione, imidazolidine-2,4-dione,pyrrolidine, tetrahydroquinolinyl, decahydroquinolinyl,tetrahydrobenzooxazepinyl dihydrodibenzooxepin and the like.

“Heteroaryl” refers to a cyclic or polycyclic aromatic radical thatcontain from one to five heteroatoms selected from N, O, and S, whereinthe nitrogen and sulfur atoms are optionally oxidized, and the nitrogenatom(s) are optionally quaternized. A heteroaryl group can be attachedto the remainder of the molecule through a heteroatom or through acarbon atom and can contain 5 to 10 carbon atoms. Non-limiting examplesof heteroaryl groups include 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl,1-pyrazolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl,2-oxazolyl, 4-oxazolyl, 5-oxazolyl, 3-isoxazolyl, 4-isoxazolyl,5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl,2-thienyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidyl and4-pyrimidyl. If not specifically stated, substituents for each of theabove noted aryl and heteroaryl ring systems are selected from the groupof acceptable substituents described herein. “Substituted heteroaryl”refers to a unsubstituted heteroaryl group as defined above in which oneor more of the ring members is bonded to a non-hydrogen atom such asdescribed above with respect to substituted alkyl groups and substitutedaryl groups. Representative substituents include straight and branchedchain alkyl groups-CH₃, —C₂H₅, —CH₂OH, —OH, —OCH₃, —OC₂H₅, —OCF₃,—OC(═O)CH₃, —OC(═O)NH₂, —OC(═O)N(CH₃)₂, —CN, —NO₂, —C(═O)CH₃, —CO₂H,—CO₂CH₃, —CONH₂, —NH₂, —N(CH₃)₂, —NHSO₂CH₃, —NHCOCH₃, —NHC(═O)OCH₃,—NHSO₂CH₃, —SO₂CH₃, —SO₂NH₂ and halo.

“Bicyclic heteroaryl” refers to bicyclic aromatic radical that containfrom one to five heteroatoms selected from N, O, and S, wherein thenitrogen and sulfur atoms are optionally oxidized, and the nitrogenatom(s) are optionally quaternized. A bicyclic heteroaryl group can beattached to the remainder of the molecule through a heteroatom orthrough a carbon atom and can contain 5 to 10 carbon atoms. Non-limitingexamples of bicyclic heteroaryl groups include 5-benzothiazolyl,purinyl, 2-benzimidazolyl, benzopyrazolyl, 5-indolyl, azaindole,1-isoquinolyl, 5-isoquinolyl, 2-quinoxalinyl, 5-quinoxalinyl, 3-quinolyland 6-quinolyl. If not specifically stated, substituents for each of theabove noted aryl and heteroaryl ring systems are selected from the groupof acceptable substituents described herein.

In each of the above embodiments designating a number of atoms e.g.“C₁₋₈” is meant to include all possible embodiments that have one feweratom. Non-limiting examples include C₁₋₇/C₂₋₈/C²⁻⁷, C₃₋₈, C₃₋₇ and thelike.

Each of the terms herein (e.g., “alkyl,” “heteroalkyl,” “aryl” and“heteroaryl”) is meant to include both “unsubstituted” and optionally“substituted” forms of the indicated radical, unless otherwiseindicated. Typically each radical is substituted with 0, 1, 2 3 4 or 5substituents, unless otherwise indicated. Examples of substituents foreach type of radical are provided below.

“Substituted” refers to a group as defined herein in which one or morebonds to a carbon(s) or hydrogen(s) are replaced by a bond tonon-hydrogen and non-carbon atom “substituents” such as, but not limitedto, a halogen atom such as F, Cl, Br, and I; an oxygen atom in groupssuch as hydroxyl groups, alkoxy groups, aryloxy, and acyloxy groups; asulfur atom in groups such as thiol groups, alkyl and aryl sulfidegroups, sulfone groups, sulfonyl groups, and sulfoxide groups; anitrogen atom in groups such as amino, alkylamines, dialkylamines,arylamines, alkylarylamines, diarylamines, alkoxyamino, hydroxyamino,acylamino, sulfonylamino, N-oxides, imides, and enamines; and otherheteroatoms in various other groups. “Substituents” also include groupsin which one or more bonds to a carbon(s) or hydrogen(s) atom isreplaced by a higher-order bond (e.g., a double- or triple-bond) to aheteroatom such as oxygen in oxo, acyl, amido, alkoxycarbonyl,aminocarbonyl, carboxyl, and ester groups; nitrogen in groups such asimines, oximes, hydrazones, and nitriles. “Substituents” further includegroups in which one or more bonds to a carbon(s) or hydrogen(s) atoms isreplaced by a bond to a cycloalkyl, heterocyclyl, aryl, and heteroarylgroups. Representative “substituents” include, among others, groups inwhich one or more bonds to a carbon or hydrogen atom is/are replaced byone or more bonds to fluoro, chloro, or bromo group. Anotherrepresentative “substituent” is the trifluoromethyl group and othergroups that contain the trifluoromethyl group. Other representative“substituents” include those in which one or more bonds to a carbon orhydrogen atom is replaced by a bond to an oxygen atom such that thesubstituted alkyl group contains a hydroxyl, alkoxy, or aryloxy group.Other representative “substituents” include alkyl groups that have anamine, or a substituted or unsubstituted alkylamine, dialkylamine,arylamine, (alkyl)(aryl)amine, diarylamine, heterocyclylamine,diheterocyclylamine, (alkyl)(heterocyclyl)amine, or(aryl)(heterocyclyl)amine group. Still other representative“substituents” include those in which one or more bonds to a carbon(s)or hydrogen(s) atoms is replaced by a bond to an alkyl, cycloalkyl,aryl, heteroaryl, or heterocyclyl group.

The herein-defined groups may include prefixes and/or suffixes that arecommonly used in the art to create additional well-recognizedsubstituent groups. As examples, “alkylamino” refers to a group of theformula —NR^(a)R^(b). Unless stated otherwise, for the following groupscontaining R^(a), R^(b), R^(c), R^(d) and R^(e): R^(a), and R^(b) areeach independently selected from H, alkyl, alkoxy, thioalkoxy,cycloalkyl, aryl, heteroaryl, or heterocyclyl or are optionally joinedtogether with the atom(s) to which they are attached to form a cyclicgroup. When R^(a) and R^(b) are attached to the same nitrogen atom, theycan be combined with the nitrogen atom to form a 5-, 6- or 7-memberedring. For example, —NR^(a)R^(b) is meant to include 1-pyrrolidinyl and4-morpholinyl.

R^(c), R^(d), R^(e) and R^(f) are each independently selected fromalkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl,heterocyclyl or alkylenearyl as defined herein.

Typically, a particular radical will have 0, 1, 2 or 3 substituents,with those groups having two or fewer substituents being preferred inthe present invention. More preferably, a radical will be unsubstitutedor monosubstituted. Most preferably, a radical will be unsubstituted.

“Substituents” for the alkyl and heteroalkyl radicals (as well as thosegroups referred to as alkylene, alkenyl, heteroalkylene, heteroalkenyl,alkynyl, cycloalkyl, heterocyclyl) can be a variety of groups selectedfrom: —OR^(a), ═O, ═NR^(a), ═N—OR^(a), —NR^(a)R^(b), —SR^(a), halogen,—SiR^(a)R^(b)R^(c), —OC(O)R^(a), —C(O)R^(a), —CO₂R^(a), —CONR^(a)R^(b),—OC(O)NR^(a)R^(b), —NR^(b)C(O)R^(a), —NR^(a)—C(O)NR^(b)R^(c),—NR^(a)—SO₂NR^(b)R^(c), —NR^(b)CO₂R^(a), —NH—C(NH₂)═NH,—NR^(a)C(NH₂)═NH, —NH—C(NH₂)═NR^(a), —S(O)R^(a), —SO₂R^(a),—SO₂NR^(a)R^(b), —NR^(b)SO₂R, —CN and —NO₂, in a number ranging fromzero to three, with those groups having zero, one or two substituentsbeing particularly preferred.

In some embodiments, “substituents”for the alkyl and heteroalkylradicals are selected from: —OR^(a), ═O, —NR^(a)R^(b), —SR^(a), halogen,—SiR^(a)R^(b)R^(c), —OC(O)R^(a), —C(O)R^(a), —CO₂R^(a), —CONR^(a)R^(b),—OC(O)NR^(a)R^(b), —NR^(b)C(O)R^(a), —NR^(b)CO₂R^(a),—NR^(a)—SO₂NR^(b)R^(c), —S(O)R^(a), —SO₂R^(a), —SO₂NR^(a)R^(b),—NR^(c)SO₂R, —CN and —NO₂, where R^(a) and R^(b) are as defined above.In some embodiments, substituents are selected from: —OR^(a), ═O,—NR^(a)R^(b), halogen, —OC(O)R^(a), —CO₂R^(a), —CONR^(a)R^(b),—OC(O)NR^(a)R^(b), —NR^(b)C(O)R^(a), —NR^(b)CO₂R^(a),—NR^(a)—SO₂NR^(b)R^(c), —SO₂R^(a), —SO₂NR^(a)R^(b), —NR″SO₂R, —CN and—NO₂.

Examples of substituted alkyl are: —(CH₂)₃NH₂, —(CH₂)₃NH(CH₃),—(CH₂)₃NH(CH₃)₂, —CH₂C(═CH₂)CH₂NH₂, —CH₂C(═O)CH₂NH₂, —CH₂S(═O)₂CH₃,—CH₂OCH₂NH₂, —CO₂H. Examples of substituents of substituted alkyl are:CH₂OH, —OH, —OCH₃, —OC₂H₅, —OCF₃, —OC(═O)CH₃, —OC(═O)NH₂,—OC(═O)N(CH₃)₂, —CN, —NO₂, —C(═O)CH₃, —CO₂H, —CO₂CH₃, —CONH₂, —NH₂,—N(CH₃)₂, —NHSO₂CH₃, —NHCOCH₃, —NHC(═O)OCH₃, —NHSO₂CH₃, —SO₂CH₃,—SO₂NH₂, and halo.

Similarly, “substituents” for the aryl and heteroaryl groups are variedand are selected from: -halogen, —OR^(a), —OC(O)R^(a), —NR^(a)R^(b),—SR^(a), —R^(a), —CN, —NO₂, —CO₂R^(a), —CONR^(a)R^(b), —C(O)R^(a),—OC(O)NR^(a)R^(b), —NR^(b)C(O)R^(a), —NR^(b)C(O)₂R^(a),—NR^(a)—C(O)NR^(b)R^(c), —NH—C(NH₂)═NH, —NR^(a)C(NH₂)═NH,—NH—C(NH₂)═NR^(a), —S(O)R^(a), —S(O)₂R^(a), —S(O)₂NR^(a)R^(b), —N₃,—CH(Ph)₂, perfluoroC₁₋₈alkoxy, and perfluoroC₁₋₈alkyl, in a numberranging from zero to the total number of open valences on the aromaticring system; and where R^(a), R^(b) and R^(c) are independently selectedfrom hydrogen, C₁₋₆alkyl and heteroalkyl, unsubstituted aryl andheteroaryl, (unsubstituted aryl)-C₁₋₈alkyl, and (unsubstitutedaryl)oxy-C₁₋₈alkyl.

Two of the “substituents” on adjacent atoms of the aryl or heteroarylring may optionally be replaced with a substituent of the formula-T-C(O)—(CH₂)q-U-, wherein T and U are independently —NH—, —O—, —CH²⁻ ora single bond, and q is 0, 1 or 2. Alternatively, two of thesubstituents on adjacent atoms of the aryl or heteroaryl ring mayoptionally be replaced with a substituent of the formula-A-(CH₂)_(r−)B—, wherein A and B are independently —CH²⁻, —O—, —NH—,—S—, —S(O)—, —S(O)²⁻, —S(O)₂NR^(a)— or a single bond, and r is 1, 2 or3. One of the single bonds of the new ring so formed may optionally bereplaced with a double bond. Alternatively, two of the substituents onadjacent atoms of the aryl or heteroaryl ring may optionally be replacedwith a substituent of the formula —(CH₂)_(S—)X—(CH₂)_(t−)—, where s andt are independently integers of from 0 to 3, and X is —O—, —NR^(a)—,—S—, —S(O)—, —S(O)²⁻, or —S(O)₂NR^(a)—. The substituent R^(a) in—NR^(a)— and —S(O)₂NR^(a)— is selected from hydrogen or unsubstitutedC₁₋₆alkyl. Otherwise, R′ is as defined above.

Unless indicated otherwise, the nomenclature of substituents that arenot explicitly defined herein are arrived at by naming the terminalportion of the functionality followed by the adjacent functionalitytoward the point of attachment. For example, the substituent“arylalkyloxycarbonyl” refers to the group (aryl)-(alkyl)-O—C(O)—.

The term “acyl” refers to the group —C(═O)R^(c) where R^(c) is alkyl,alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl orheterocyclyl. Acyl includes the “acetyl” group—C(═O)CH₃.

“Acylamino-” refers to the group —NR^(a)C(═O)R^(c) where R^(c) is alkyl,alkenyl, alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl orheterocyclyl.

“Acyloxy” refers to —OC(═O)—R^(c) where R^(c) is alkyl, alkenyl,alkynyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl or heterocyclyl.

“Alkoxy” refers to —OR^(d) wherein R^(d) is alkyl as defined herein.Representative examples of alkoxy groups include methoxy, ethoxy,t-butoxy, trifluoromethoxy, and the like.

“Alkoxyamino” refers to the group —NHOR^(d) where R^(d) is alkyl.

“Alkoxyalkyleneamino” refers to the group —NR^(a)-alkylene-OR^(d) whereR^(d) is alkyl and —NR^(a)— is defined in amino.

“Alkoxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) is alkyl.Representative alkoxycarbonyl groups include, for example, those shownbelow.

These alkoxycarbonyl groups can be further substituted as will beapparent to those having skill in the organic and medicinal chemistryarts in conjunction with the disclosure herein.

“Alkoxycarbonylalkylene” refers to the group -alkylene-C(═O)OR^(d)wherein R^(d) is alkyl.

“Alkoxycarbonylamino” refers to to —NR^(a)C(═O)OR^(d) wherein R^(d) isalkyl.

“Alkoxycarbonylaminoalkylene” refers to to -alkylene-NR^(a)C(═O)OR^(d)wherein R^(d) is alkyl.

“Alkoxycarbonylalkyleneaminosulfonyl” refers to to —SO₂NR^(a)-alkyleneC(═O)OR^(d) wherein R^(d) is alkyl.

“Alkoxysulfonylamino” refers to the group —NR^(a)S(═O)₂—OR^(d) whereR^(d) is alkyl.

“Alkylcarbonyl” refers to the group —C(═O)R^(c) where R^(c) is alkyl.

“Alkylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) is alkyl.

“Alkylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) is alkyl.Representative alkylcarbonylamino groups include, for example,—NHC(═O)CH₃, —NHC(═O)CH₂CH₃, —NHC(═O)CH₂NH(CH₃), —NHC(═O)CH₂N(CH₃)₂, or—NHC(═O)(CH₂)₃OH.

“Alkylheterocyclyl” refers to the group -heterocyclyl-R^(d). where R^(d)is alkyl.

“Alkylheterocyclylalkylene” refers to the group-alkylene-heterocyclyl-R^(d). where R^(d) is alkyl.

“Alkylsulfanyl”, “alkylthio”, or “thioalkoxy” refers to the groupS—R^(d). where R^(d) is alkyl.

“Alkylsulfinyl” refers to —S(═O)R^(c) where R^(c) is alkyl.Alkylsulfonyl groups employed in compounds of the present invention aretypically C₁₋₆alkylsulfinyl groups.

“Alkylsulfonyl” refers to —S(═O)₂R^(c) where R^(c) is alkyl.Alkylsulfonyl groups employed in compounds of the present invention aretypically C₁₋₆alkylsulfonyl groups.

“Alkylsulfonylalkylene” refers to -alkylene-S(═O)₂R^(c) where R^(c) isalkyl. Alkylsulfonyl groups employed in compounds of the presentinvention are typically C₁₋₆alkylsulfonyl groups.

“Alkylsulfonylamino” refers to —NR^(a)S(═O)₂—R^(e) wherein R^(e) isalkyl.

“Alkynyloxy” refers to the group —O-alkynyl, wherein alkynyl is asdefined herein. Alkynyloxy includes, by way of example, ethynyloxy,propynyloxy, and the like.

“Amidino” refers to the group —C(═NR^(a))NR^(b)R^(c), wherein R^(b) andR^(c) independently are selected from the group consisting of hydrogen,alkyl, substituted alkyl, alkenyl, alkynyl, aryl, cycloalkyl,cycloalkenyl, heteroaryl, heterocyclic, and where R^(b) and R^(c) areoptionally joined together with the nitrogen bound thereto to form aheterocyclic or substituted heterocyclic group. R^(a) is selected fromthe group consisting of hydrogen, alkyl, alkenyl, alkynyl, cycloalkynyl,aryl, cycloalkyl, cycloalkenyl, heteroaryl, heterocyclic, substitutedheterocyclic, nitro, nitroso, hydroxy, alkoxy, cyano, —N═N—N-alkyl,—N(alkyl)SO₂-alkyl, —N═N=N-alkyl, acyl and —SO₂-alkyl.

“Amino” refers to a monovalent radical —NR^(a)R^(b) or divalent radical—NR^(a)—. The term includes “alkylamino” which refers to the group—NR^(a)R^(b) where R^(a) is alkyl and R^(b) is H or alkyl. The term alsoincludes “arylamino” which refers to the group NR^(a)R^(b) where atleast one R^(a) or R^(b) is aryl. The term also includes“(alkyl)(aryl)amino” which refers to the group —NR^(a)R^(b) where R^(a)is alkyl and R^(b) is aryl. Additionally, for dialkylamino groups, thealkyl portions can be the same or different and can also be combined toform a 3-7 membered ring with the nitrogen atom to which each isattached. Accordingly, a group represented as —R^(a)R^(b) is meant toinclude piperidinyl, pyrrolidinyl, morpholinyl, azetidinyl and the like.

“Aminoalkoxy” refers to —O-alkylene-NR^(a)R^(b).

“Aminoalkylene” refers to -alkylene-NR^(a)R^(b).

“Aminoalkylenecarbonyl” refers to —C(═O)-alkylene-NR^(a)R^(b).

“Aminoalkyleneaminocarbonyl” refers to—C(═O)NR^(a)-alkylene-NR^(a)R^(b).

“Aminoaryl” refers to -aryl-NR^(a)R^(b).

“Aminocarbonyl” or “aminoacyl” refers to the amide —C(═O)—NR^(a)R^(b).The term “alkylaminocarbonyl” refers herein to the group—C(═O)—NR^(a)R^(b) where R^(a) is alkyl and R^(b) is H or alkyl. Theterm “arylaminocarbonyl” refers herein to the group —C(═O)—NR^(a)R^(b)where R^(a) or R^(b) is aryl. Representative aminocarbonyl groupsinclude, for example, those shown below. These aminocarbonyl group canbe further substituted as will be apparent to those having skill in theorganic and medicinal chemistry arts in conjunction with the disclosureherein.

“Aminocarbonylalkoxy” refers to —O-alkylene-C(═O)—NR^(a)R^(b) whereinR^(a) is hydrogen or alkyl and R^(a) and R^(b) independently areselected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl,aryl, cycloalkyl, cycloalkenyl, heteroaryl, heterocyclic, and whereR^(a) and R^(b) are optionally joined together with the nitrogen boundthereto to form a heterocyclic or substituted heterocyclic group.

“Aminocarbonylalkylene” refers to -alkylene-C(═O)—NR^(a)R^(b) whereinR^(a) is hydrogen or alkyl and R^(a) and R^(b) independently areselected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl,aryl, cycloalkyl, cycloalkenyl, heteroaryl, heterocyclic, and whereR^(a) and R^(b) are optionally joined together with the nitrogen boundthereto to form a heterocyclic or substituted heterocyclic group.

“Aminocarbonylalkyleneaminosulfonyl” refers to—S(O)₂NR^(a)-alkylene-C(═O)—NR^(a)R^(b) wherein each R^(a) is hydrogenor alkyl and R^(a) and R^(b) independently are selected from the groupconsisting of hydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl,cycloalkenyl, heteroaryl, heterocyclic, and where R^(a) and R^(b) of theamino group are optionally joined together with the nitrogen boundthereto to form a heterocyclic or substituted heterocyclic group.

“Aminocarbonylamino” refers to the group —NR^(a)C(O)NR^(a)R^(b), whereinR^(a) is hydrogen or alkyl and R^(a) and R^(b) independently areselected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl,aryl, cycloalkyl, cycloalkenyl, heteroaryl, heterocyclic, and whereR^(a) and R^(b) are optionally joined together with the nitrogen boundthereto to form a heterocyclic or substituted heterocyclic group.

“Aminocarbonylaminoalkylene” refers to the group-alkylene-NR^(a)C(O)NR^(a)R^(b), wherein R^(a) is hydrogen or alkyl andR^(a) and R^(b) independently are selected from the group consisting ofhydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, cycloalkenyl,heteroaryl, heterocyclic, and where R^(a) and R^(b) are optionallyjoined together with the nitrogen bound thereto to form a heterocyclicor substituted heterocyclic group.

“Aminocarboxyalkylene” refers to the group -alkylene-OC(O)NR^(a)R^(b),wherein R^(a) is hydrogen or alkyl and R^(a) and R^(b) independently areselected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl,aryl, cycloalkyl, cycloalkenyl, heteroaryl, heterocyclic, and whereR^(a) and R^(b) are optionally joined together with the nitrogen boundthereto to form a heterocyclic or substituted heterocyclic group.

“Aminosulfonyl” refers to —S(O)₂NR^(a)R^(b) where R is independently areselected from the group consisting of hydrogen, alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl,substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl,substituted cycloalkenyl, heteroaryl, substituted heteroaryl,heterocyclic, substituted heterocyclic and where R^(a) and R^(b) areoptionally joined together with the nitrogen bound thereto to form aheterocyclic or substituted heterocyclic group and alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl,cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Aminosulfonylalkylene” refers to -alkylene-S(O)₂NR^(a)R^(b) where R isindependently are selected from the group consisting of hydrogen, alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl,cycloalkenyl, substituted cycloalkenyl, heteroaryl, substitutedheteroaryl, heterocyclic, substituted heterocyclic and where R^(a) andR^(b) are optionally joined together with the nitrogen bound thereto toform a heterocyclic or substituted heterocyclic group and alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

The term “alkylaminosulfonyl” refers herein to the group—S(O)₂NR^(a)R^(b) where R^(a) is alkyl and R^(b) is H or alkyl. The term“alkylarylsulfonyl” refers herein to the group—S(O)₂NR^(a)R^(b) whereR^(a) or R^(b) is alkylaryl.

“Aminosulfonyloxy” refers to the group —O—SO₂NR^(a)R^(b), wherein R^(a)and R^(b) independently are selected from the group consisting ofhydrogen, alkyl, alkenyl, alkynyl, aryl, cycloalkyl, cycloalkenyl,heteroaryl and heterocyclic; R^(a) and R^(b) are optionally joinedtogether with the nitrogen bound thereto to form a heterocyclic orsubstituted heterocyclic group.

“Aminosulfonylamino” refers to the group —NR^(a)—SO₂NR^(b)R^(c), whereinR^(a) is hydrogen or alkyl and R^(b) and R^(c) independently areselected from the group consisting of hydrogen, alkyl, substitutedalkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl,substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl,substituted cycloalkenyl, heteroaryl, substituted heteroaryl,heterocyclic, and substituted heterocyclic and where R^(b) and R^(c) areoptionally joined together with the nitrogen bound thereto to form aheterocyclic or substituted heterocyclic group, and wherein alkyl,substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substitutedalkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, aryl, substituted aryl, heteroaryl, substitutedheteroaryl, heterocyclic and substituted heterocyclic are as definedherein.

“Aminothiocarbonyl” refers to the group —C(S)NR^(a)R^(b), wherein R^(a)and R^(b) independently are selected from the group consisting ofhydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl,alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl,substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl,heteroaryl, substituted heteroaryl, heterocyclic, and substitutedheterocyclic and where R^(a) and R^(b) are optionally joined togetherwith the nitrogen bound thereto to form a heterocyclic or substitutedheterocyclic group, and wherein alkyl, substituted alkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl,substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl,substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, andsubstituted heterocyclic are as defined herein.

“Aminothiocarbonylamino” refers to the group—NR^(a)C(S)NR^(a)R^(b),wherein R^(a) is hydrogen or alkyl and R^(b) and R^(c) are optionallyjoined together with the nitrogen bound thereto to form a heterocyclicor substituted heterocyclic group.

“Arylalkoxycarbonylamino” refers to the group—NR^(a)C(═O)O-alkylene-R^(c) where R^(c) is aryl.

“Arylcarbonyl” refers to the group —C(═O)R^(c) where R^(c) is aryl.

“Arylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) is aryl.

“Arylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) is aryl.

“Aryloxy” refers to —OR^(d) where R^(d) is aryl. Representative examplesof aryloxy groups include phenoxy, naphthoxy, and the like.

“Aryloxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) is aryl.

“Aryloxycarbonylamino” refers to —NR^(a)C(═O)OR^(d) wherein R^(d) isaryl.

“Arylsulfanyl”, “arylthio”, or “thioaryloxy” refers to the groupS—R^(d). where R^(d) is aryl.

“Arylsulfonyl” refers to —S(═O)₂R^(e) where R^(e) is aryl.

“Arylsulfonylamino” refers to —NR^(a)S(═O)₂—R^(e) wherein R^(e) is aryl.

“Arylthio” refers to the group —S-aryl, wherein aryl is as definedherein. In other embodiments, sulfur may be oxidized to —S(O)— or —SO₂—moieties. The sulfoxide may exist as one or more stereoisomers.

“Azido” refers to —N₃.

“Bond” when used a element in a Markush group means that thecorresponding group does not exist, and the groups of both sides aredirectly linked.

“Carbonyl” refers to the divalent group —C(═O)—.

“Carboxy” or “carboxyl” refers to the group —CO₂H.

“Carboxyalkylene” refers to the group -alkylene-CO₂H.

“Carboxyalkylenesulfonylamino” refers to the group—NR^(a)SO₂-alkylene-CO₂H.

“Carboxyl ester”, “carbonylalkoxy” or “carboxy ester” refers to thegroup —C(═O)OR^(c).

“(Carboxyl ester)amino” refers to the groups —NR^(a)—C(O)OR^(c), whereR^(a) is alkyl or hydrogen.

“(Carboxyl ester)oxy” or “Carbonate ester” refers to the groups—O—C(═O)OR^(c).

“Cyano” refers to —CN.

“Cyanoalkylenecarbonyl” refers to —C(═O)-alkylene-CN.

“Cycloalkoxy” refers to —OR^(d) where R^(d) is cycloalkyl.

“Cycloalkoxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) iscycloalkyl.

“Cycloalkoxycarbonylamino” refers to to —NR^(a)C(═O)OR^(d) wherein R^(d)is cycloalkyl.

“Cycloalkylalkylene” refers to a radical -R^(x)R^(y) wherein R^(x) is analkylene group and R^(y) is a cycloalkyl group as defined herein, e.g.,cyclopropylmethyl, cyclohexenylpropyl, 3-cyclohexyl-2-methylpropyl, andthe like.

“Cycloalkylcarbonyl” refers to the group —C(═O)R^(c) where R^(c) iscycloalkyl.

“Cycloalkylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) iscycloalkyl.

“Cycloalkylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) iscycloalkyl.

“Cycloalkylsulfonylamino” refers to —NR^(a)S(═O)₂—R^(c) wherein R^(c) iscycloalkyl.

“Cycloalkylthio” refers to —S-cycloalkyl. In other embodiments, sulfurmay be oxidized to —S(O)— or —SO₂— moieties. The sulfoxide may exist asone or more stereoisomers.

“Cycloalkenylox” refers to —O-cycloalkenyl.

“Cycloalkenylthio” refers to —S-cycloalkenyl. In other embodiments,sulfur may be oxidized to sulfinyl or sulfonyl moieties. The sulfoxidemay exist as one or more stereoisomers.

“Ester” refers to —C(═O)OR^(d) wherein R^(d) is alkyl, cycloalkyl, aryl,heteroaryl, or heterocyclyl.

“Guanidino” refers to the group —NHC(═NH)NH₂.

“Halo” or “halogen” by themselves or as part of another substituent,mean, unless otherwise stated, a fluorine, chlorine, bromine, or iodineatom. Additionally, terms such as “haloalkyl”, are meant to includealkyl in which one or more hydrogen is substituted with halogen atomswhich can be the same or different, in a number ranging from one up tothe maximum number of halogens permitted e.g. for alkyl, (2m′+1), wherem′ is the total number of carbon atoms in the alkyl group. For example,the term “haloC₁₋₈alkyl” is meant to include trifluoromethyl,2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like. Theterm “perhaloalkyl” means, unless otherwise stated, alkyl substitutedwith (2m′+1) halogen atoms, where m′ is the total number of carbon atomsin the alkyl group. For example, the term “perhaloC₁₋₈ alkyl”, is meantto include trifluoromethyl, pentachloroethyl,1,1,1-trifluoro-2-bromo-2-chloroethyl, and the like. Additionally, term“haloalkoxy” refers to an alkoxy radical substituted with one or morehalogen atoms.

“Heteroalkyl” means an alkyl radical as defined herein with one, two orthree substituents independently selected from cyano, —OR^(w),—NR^(x)R^(y), and —S(O)_(n)R^(z) (where n is an integer from 0 to 2),with the understanding that the point of attachment of the heteroalkylradical is through a carbon atom of the heteroalkyl radical. R^(w) ishydrogen, alkyl, cycloalkyl, cycloalkyl-alkyl, aryl, araalkyl,alkoxycarbonyl, aryloxycarbonyl, carboxamido, or mono- ordi-alkylcarbamoyl. R^(x) is hydrogen, alkyl, cycloalkyl,cycloalkyl-alkyl, aryl or araalkyl. Ry is hydrogen, alkyl, cycloalkyl,cycloalkyl-alkyl, aryl, araalkyl, alkoxycarbonyl, aryloxycarbonyl,carboxamido, mono- or di-alkylcarbamoyl or alkylsulfonyl. R^(z) ishydrogen (provided that n is 0), alkyl, cycloalkyl, cycloalkyl-alkyl,aryl, araalkyl, amino, mono-alkylamino, di-alkylamino, or hydroxyalkyl.Representative examples include, for example, 2-hydroxyethyl,2,3-dihydroxypropyl, 2-methoxyethyl, benzyloxymethyl, 2-cyanoethyl, and2-methylsulfonyl-ethyl. For each of the above, R^(w), R^(x), R^(y), andR^(z) can be further substituted by amino, fluorine, alkylamino,di-alkylamino, OH or alkoxy. Additionally, the prefix indicating thenumber of carbon atoms (e.g., C₁-C₁₀) refers to the total number ofcarbon atoms in the portion of the heteroalkyl group exclusive of thecyano, —OR^(w), —NR^(x)R^(y), or —S(O)_(n)R^(z) portions.

“Heteroarylalkenyl” refers to the group -alkenyl-R^(c) where R^(c) isheteroaryl.

“Heteroarylcarbonyl” refers to the group —C(═O)R^(r) where R^(e) isheteroaryl.

“Heteroarylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) isheteroaryl.

“Heteroarylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) isheteroaryl.

“Heteroaryloxy” refers to —OR^(d) where R^(d) is heteroaryl.

“Heteroaryloxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) isheteroaryl.

“Heteroaryloxycarbonylamino” refers to to —NR^(a)C(═O)OR^(d) whereinR^(d) is heteroaryl.

“Heteroarylsulfonyllamino” refers to —NR^(a)S(═O)₂—R^(e) wherein R^(e)is heteroaryl.

“Heteroarylthio” refers to the group —S-heteroaryl. In otherembodiments, sulfur may be oxidized to —S(O)— or —SO₂— moieties. Thesulfoxide may exist as one or more stereoisomers.

“Heterocyclylalkyl” or “Cycloheteroalkyl-alkyl” means a radical-R^(x)R^(y) where R^(x) is an alkylene group and R^(y) is a heterocyclylgroup as defined herein, e.g., tetrahydropyran-2-ylmethyl,4-(4-substituted-phenyl)piperazin-1-ylmethyl, 3-piperidinylethyl, andthe like.

“Heterocycloxycarbonylamino” refers to to —NR^(a)C(═O)OR^(d) whereinR^(d) is heterocyclyl.

“Heterocyclylcarbonyl” refers to the —C(═O)R^(c) where R^(c) isheterocyclyl.

“Heterocyclylcarbonylamino” refers to —NR^(a)C(═O)R^(c) wherein R^(c) isheterocyclyl.

“Heterocyclylcarbonyloxy” refers to —OC(═O)—R^(c) where R^(c) sheterocyclyl.

“Heterocyclyloxy” refers to —OR^(d) where R^(d) is heterocyclyl.

“Heterocyclyloxycarbonyl” refers to —C(═O)OR^(d) wherein R^(d) isheterocyclyl.

“Heterocyclylsulfonyl” refers to —S(═O)₂R^(e) where R^(e) isheterocyclyl.

“Heterocyclylsulfonyllamino” refers to —NR^(a)S(═O)₂—R^(e) wherein R^(e)is heterocyclyl.

“Heterocyclylthio” refers to the group —S-heterocycyl. In otherembodiments, sulfur may be oxidized to —S(O)— or —SO₂— moieties. Thesulfoxide may exist as one or more stereoisomers.

“Hydroxy” or “hydroxyl” refers to the group —OH.

“Hydroxyalkylene” refers to the group -alkylene-OH.

“Hydroxyalkyleneamino” refers to the group —NR^(a)-alkylene-OH.

“Hydroxyalkyleneaminocarbonyl” refers to the group—C(═O)NR^(a)-alkylene-OH.

“Hydroxyalkyleneaminosulfonyl” refers to the group—SO₂NR^(a)-alkylene-OH.

“Hydroxyamino” refers to the group —NHOH.

“Hydroxyalkylenecarbonylamino” refers to the group—NR^(a)C(═O)-alkylene-OH.

“Imino” refers to the group ═NR^(a).

“Nitro” refers to —NO₂.

“Nitroso” refers to the group—NO.

The terms “optional” or “optionally” as used throughout thespecification means that the subsequently described event orcircumstance may but need not occur, and that the description includesinstances where the event or circumstance occurs and instances in whichit does not. For example, “heterocyclo group optionally mono- ordi-substituted with an alkyl group means that the alkyl may but need notbe present, and the description includes situations where theheterocyclo group is mono- or disubstituted with an alkyl group andsituations where the heterocyclo group is not substituted with the alkylgroup.

“Optionally substituted” means a ring which is optionally substitutedindependently with substituents. A site of a group that is unsubstitutedmay be substituted with hydrogen.

“Oxo” refers to the divalent group ═O.

“Sulfanyl” refers to the group —SR^(f) where R^(f) is as defined herein.

“Sulfinyl” refers to the group —S(═O)—R^(e) where R^(e) is as definedherein.

“Sulfonic acid” refers to the group —S(O)₂—OH.

“Sulfonyl” refers to the group —S(O)₂—R^(e) where R^(e) is as definedherein.

“Sulfonylamino” refers to —NR^(a)S(═O)₂—R^(e) where R^(a) is selectedfrom the group consisting of hydrogen, alkyl, alkenyl, alkynyl, aryl,cycloalkyl, cycloalkenyl, heteroaryl and heterocyclyl and R^(e) is asdefined herein.

“Sulfonyloxy” refers to the group —OSO₂—R^(c).

Compounds that have the same molecular formula but differ in the natureor sequence of bonding of their atoms or the arrangement of their atomsin space are termed “isomers”. Isomers that differ in the arrangement oftheir atoms in space are termed “stereoisomers”. “Stereoisomer” and“stereoisomers” refer to compounds that exist in differentstereoisomeric forms if they possess one or more asymmetric centers or adouble bond with asymmetric substitution and, therefore, can be producedas individual stereoisomers or as mixtures. Stereoisomers includeenantiomers and diastereomers. Stereoisomers that are not mirror imagesof one another are termed “diastereomers” and those that arenon-superimposable mirror images of each other are termed “enantiomers”.When a compound has an asymmetric center, for example, it is bonded tofour different groups, a pair of enantiomers is possible. An enantiomercan be characterized by the absolute configuration of its asymmetriccenter and is described by the R- and S-sequencing rules of Cahn andPrelog, or by the manner in which the molecule rotates the plane ofpolarized light and designated as dextrorotatory or levorotatory (i.e.,as (+) or (−)-isomers respectively). A chiral compound can exist aseither individual enantiomer or as a mixture thereof. A mixturecontaining equal proportions of the enantiomers is called a “racemicmixture”. Unless otherwise indicated, the description is intended toinclude individual stereoisomers as well as mixtures. The methods forthe determination of stereochemistry and the separation of stereoisomersare well-known in the art (see discussion in Chapter 4 of ADVANCEDORGANIC CHEMISTRY, 4th edition J. March, John Wiley and Sons, New York,1992) differ in the chirality of one or more stereocenters.

“Thioacyl” refers to the groups R^(a)—C(S)—.

“Thiol” refers to the group —SH.

“Tautomer” refers to alternate forms of a molecule that differ in theposition of a proton, such as enol-keto and imine-enamine tautomers, orthe tautomeric forms of heteroaryl groups containing a —N═C(H)—NH— ringatom arrangement, such as pyrazoles, imidazoles, benzimidazoles,triazoles, and tetrazoles. A person of ordinary skill in the art wouldrecognize that other tautomeric ring atom arrangements are possible.

It is understood that in all substituted groups defined above, polymersarrived at by defining substituents with further substituents tothemselves (e.g., substituted aryl having a substituted aryl group as asubstituent which is itself substituted with a substituted aryl group,which is further substituted by a substituted aryl group, etc.) are notintended for inclusion herein. In such cases, the maximum number of suchsubstitutions is three. For example, serial substitutions of substitutedaryl groups are limited to -substituted aryl-(substitutedaryl)-substituted aryl.

“Protecting group” refers to a group of atoms that, when attached to areactive functional group in a molecule, mask, reduce or prevent thereactivity of the functional group. Typically, a protecting group may beselectively removed as desired during the course of a synthesis.Examples of protecting groups can be found in Greene and Wuts,Protective Groups in Organic Chemistry, 3^(rd) Ed., 1999, John Wiley &Sons, NY and Harrison et al., Compendium of Synthetic Organic Methods,Vols. 1-8, 1971-1996, John Wiley & Sons, NY. Representative aminoprotecting groups include, but are not limited to, formyl, acetyl,trifluoroacetyl, benzyl, benzyloxycarbonyl (“CBZ”), tert-butoxycarbonyl(“Boc”), trimethylsilyl (“TMS”), 2-trimethylsilyl-ethanesulfonyl(“TES”), trityl and substituted trityl groups, allyloxycarbonyl,9-fluorenylmethyloxycarbonyl (“FMOC”), nitro-veratryloxycarbonyl(“NVOC”) and the like. Representative hydroxy protecting groups include,but are not limited to, those where the hydroxy group is either acylatedor alkylated such as benzyl and trityl ethers, as well as alkyl ethers,tetrahydropyranyl ethers, trialkylsilyl ethers (e.g., TMS or TIPPSgroups) and allyl ethers.

The term “pharmaceutically acceptable salts” is meant to include saltsof the active compounds which are prepared with relatively nontoxicacids or bases, depending on the particular substituents found on thecompounds described herein. When compounds of the present inventioncontain relatively acidic functionalities, base addition salts can beobtained by contacting the neutral form of such compounds with asufficient amount of the desired base, either neat or in a suitableinert solvent. Examples of salts derived frompharmaceutically-acceptable inorganic bases include aluminum, ammonium,calcium, copper, ferric, ferrous, lithium, magnesium, manganic,manganous, potassium, sodium, zinc and the like. Salts derived frompharmaceutically-acceptable organic bases include salts of primary,secondary and tertiary amines, including substituted amines, cyclicamines, naturally-occurring amines and the like, such as arginine,betaine, caffeine, choline, N,N′-dibenzylethylenediamine, diethylamine,2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine,ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine,glucosamine, histidine, hydrabamine, isopropylamine, lysine,methylglucamine, morpholine, piperazine, piperidine, polyamine resins,procaine, purines, theobromine, triethylamine, trimethylamine,tripropylamine, tromethamine and the like. When compounds of the presentinvention contain relatively basic functionalities, acid addition saltscan be obtained by contacting the neutral form of such compounds with asufficient amount of the desired acid, either neat or in a suitableinert solvent. Examples of pharmaceutically acceptable acid additionsalts include those derived from inorganic acids like hydrochloric,hydrobromic, nitric, carbonic, monohydrogencarbonic, phosphoric,monohydrogenphosphoric, dihydrogenphosphoric, sulfuric,monohydrogensulfuric, hydriodic, or phosphorous acids and the like, aswell as the salts derived from relatively nontoxic organic acids likeacetic, propionic, isobutyric, malonic, benzoic, succinic, suberic,fumaric, mandelic, phthalic, benzenesulfonic, p-tolylsulfonic, citric,tartaric, methanesulfonic, and the like. Also included are salts ofamino acids such as arginate and the like, and salts of organic acidslike glucuronic or galactunoric acids and the like (see, e.g., Berge, S.M. et al., “Pharmaceutical Salts,” Journal of Pharmaceutical Science,66:1-19, 1977). Certain specific compounds of the present inventioncontain both basic and acidic functionalities that allow the compoundsto be converted into either base or acid addition salts.

The neutral forms of the compounds may be regenerated by contacting thesalt with a base or acid and isolating the parent compound in theconventional manner. The parent form of the compound differs from thevarious salt forms in certain physical properties, such as solubility inpolar solvents, but otherwise the salts are equivalent to the parentform of the compound for the purposes of the present invention.

In addition to salt forms, the present invention provides compoundswhich are in a prodrug ester form. “Prodrug”s of the compounds describedherein are those compounds that readily undergo chemical changes underphysiological conditions to provide the compounds of the presentinvention. Additionally, prodrugs can be converted to the compounds ofthe present invention by chemical or biochemical methods in an ex vivoenvironment. For example, prodrugs can be slowly converted to thecompounds of the present invention when placed in a transdermal patchreservoir with a suitable enzyme or chemical reagent. Prodrugs arefrequently, but not necessarily, pharmacologically inactive untilconverted into the active drug. Prodrugs are typically obtained bymasking a functional group in the drug believed to be in part requiredfor activity with a progroup (defined below) to form a promoiety whichundergoes a transformation, such as cleavage, under the specifiedconditions of use to release the functional group, and hence the activedrug. The cleavage of the promoiety may proceed spontaneously, such asby way of a hydrolysis reaction, or it may be catalyzed or induced byanother agent, such as by an enzyme, by light, by acid or base, or by achange of or exposure to a physical or environmental parameter, such asa change of temperature. The agent may be endogenous to the conditionsof use, such as an enzyme present in the cells to which the prodrug isadministered or the acidic conditions of the stomach, or it may besupplied exogenously.

“Progroup” refers to a type of protecting group that, when used to maska functional group within an active drug to form a promoiety, convertsthe drug into a prodrug. Progroups are typically attached to thefunctional group of the drug via bonds that are cleavable underspecified conditions of use. Thus, a progroup is that portion of apromoiety that cleaves to release the functional group under thespecified conditions of use. As a specific example, an amide promoietyof the formula —NH—C(O)CH₃ comprises the progroup —C(O)CH₃.

A wide variety of progroups, as well as the resultant promoieties,suitable for masking functional groups in the active syk and/or JAKselective inhibitory compounds to yield prodrugs are well-known in theart. For example, a hydroxyl functional group may be masked as asulfonate, ester (such as acetate or maleate) or carbonate promoiety,which may be hydrolyzed in vivo to provide the hydroxyl group. An aminofunctional group may be masked as an amide, carbamate, imine, urea,phosphenyl, phosphoryl or sulfenyl promoiety, which may be hydrolyzed invivo to provide the amino group. A carboxyl group may be masked as anester (including methyl, ethyl, pivaloyloxymethyl, silyl esters andthioesters), amide or hydrazide promoiety, which may be hydrolyzed invivo to provide the carboxyl group. The invention includes those estersand acyl groups known in the art for modifying the solubility orhydrolysis characteristics for use as sustained-release or prodrugformulations. Other specific examples of suitable progroups and theirrespective promoieties will be apparent to those of skill in the art.

Certain compounds of the present invention can exist in unsolvated formsas well as solvated forms, including hydrated forms. “Solvate” refers toa complex formed by combination of solvent molecules with molecules orions of the solute. The solvent can be an organic compound, an inorganiccompound, or a mixture of both. Some examples of solvents include, butare not limited to, methanol, N,N-dimethylformamide, tetrahydrofuran,dimethylsulfoxide, and water. In general, the solvated forms areequivalent to unsolvated forms and are intended to be encompassed withinthe scope of the present invention. Certain compounds of the presentinvention may exist in multiple crystalline or amorphous forms. Ingeneral, all physical forms are equivalent for the uses contemplated bythe present invention and are intended to be within the scope of thepresent invention.

Certain compounds of the present invention possess asymmetric carbonatoms (optical centers) or double bonds; the racemates, diastereomers,geometric isomers, regioisomers and individual isomers (e.g., separateenantiomers) are all intended to be encompassed within the scope of thepresent invention. These isomers can be resolved or asymmetricallysynthesized using conventional methods to render the isomers “opticallypure”, i.e., substantially free of its other isomers. If, for instance,a particular enantiomer of a compound of the present invention isdesired, it may be prepared by asymmetric synthesis, or by derivationwith a chrial auxilliary, where the resulting diastereomeric mixture isseparated and the auxilliary group cleaved to provide the pure desiredenantiomers. Alternatively, where the molecule contains a basicfunctional group, such as amino, or an acidic functional group, such ascarboxyl, diastereomeric salts are formed with an appropriateoptically-active acid or base, followed by resolution of thediasteromers thus formed by fractional crystallization orchromatographic means well known in the art, and subsequent recovery ofthe pure enantiomers.

The compounds of the present invention may also contain unnaturalproportions of atomic isotopes at one or more of the atoms thatconstitute such compounds. For example, the compounds may beradiolabeled with radioactive isotopes, such as for example tritium(³H), iodine-125 (¹²⁵I) or carbon-14 (¹⁴C). All isotopic variations ofthe compounds of the present invention, whether radioactive or not, areintended to be encompassed within the scope of the present invention.

The term “administering” refers to oral administration, administrationas a suppository, topical contact, intravenous, intraperitoneal,intramuscular, intralesional, intranasal or subcutaneous administration,or the implantation of a slow-release device e.g., a mini-osmotic pump,to a subject. Administration is by any route, including parenteral andtransmucosal (e.g., buccal, sublingual, palatal, gingival, nasal,vaginal, rectal, or transdermal). Parenteral administration includes,e.g., intravenous, intramuscular, intra-arteriole, intradermal,subcutaneous, intraperitoneal, intraventricular, and intracranial. Othermodes of delivery include, but are not limited to, the use of liposomalformulations, intravenous infusion, transdermal patches, etc.

An “agonist” or “activator” refers to an agent or molecule that binds toa receptor of the invention, stimulates, increases, opens, activates,facilitates, enhances activation or enzymatic activity, sensitizes or upregulates the activity of a receptor of the invention.

An “antagonist” or “inhibitor” refers to an agent or molecule thatinhibits or binds to, partially or totally blocks stimulation oractivity, decreases, closes, prevents, delays activation or enzymaticactivity, inactivates, desensitizes, or down regulates the activity of areceptor of the invention. As used herein, “antagonist” also includes areverse or inverse agonist.

As used herein, the term “condition or disorder responsive to modulationof syk and/or JAK” and related terms and phrases refer to a condition ordisorder associated with inappropriate, e.g., less than or greater thannormal, activity of syk and/or JAK and at least partially responsive toor affected by modulation of syk and/or JAK (e.g., syk and/or JAKantagonist or agonist results in some improvement in patient well-beingin at least some patients). Inappropriate functional activity of sykand/or JAK might arise as the result of expression of syk and/or JAK incells which normally do not express the receptor, greater than normalproduction of syk and/or JAK, or slower than normal metabolicinactivation or elimination of syk and/or JAK or its active metabolites,increased expression of syk and/or JAK or degree of intracellularactivation (leading to, e.g., inflammatory and immune-related disordersand conditions) or decreased expression of syk and/or JAK. A conditionor disorder associated with syk and/or JAK may include a “syk and/or JAK-mediated condition or disorder”.

As used herein, the phrases “a condition or disorder mediated at leastin part by syk or JAK kinase activity”, and related phrases and termsrefer to a condition or disorder characterized by inappropriate, e.g.,greater than normal, syk and/or JAK activity. Inappropriate syk and/orJAK functional activity might arise as the result of syk and/or JAKexpression in cells which normally do not express syk and/or JAK orincreased syk and/or JAK expression or degree of intracellularactivation (leading to, e.g., inflammatory and immune-related disordersand conditions). A condition or disorder mediated at least in part bysyk or JAK kinase activity may be completely or partially mediated byinappropriate syk and/or JAK functional activity. However, a conditionor disorder mediated at least in part by syk or JAK kinase activity isone in which modulation of syk and/or JAK results in some effect on theunderlying condition or disorder (e.g., an syk and/or JAK antagonistresults in some improvement in patient well-being in at least somepatients).

The term “inflammation” as used herein refers to infiltration of whiteblood cells (e.g., leukocytes, monocytes, etc.) into the area beingtreated for restenosis.

The term “intervention” refers to an action that produces an effect orthat is intended to alter the course of a disease process. For example,“vascular intervention” refers to the use of an intravascular proceduresuch as angioplasty or a stent to open an obstructed blood vessel.

The term “intravascular device” refers to a device useful for a vascularrecanalization procedure to restore blood flow through an obstructedblood vessel. Examples of intravascular devices include, withoutlimitation, stents, balloon catheters, autologous venous/arterialgrafts, prosthetic venous/arterial grafts, vascular catheters, andvascular shunts.

As used herein, the term “JAK” refers to a Janus kinase (RefSeqAccession No. P-43408) or a variant thereof that is capable of mediatinggene expression in vitro or in vivo. JAK variants include proteinssubstantially homologous to native JAK, i.e., proteins having one ormore naturally or non-naturally occurring amino acid deletions,insertions or substitutions (e.g., JAK derivatives, homologs andfragments). The amino acid sequence of JAK variant preferably is atleast about 80% identical to a native JAK, more preferably at leastabout 90% identical, and most preferably at least about 95% identical.

The term “leukocyte” refers to any of the various blood cells that havea nucleus and cytoplasm, separate into a thin white layer when wholeblood is centrifuged, and help protect the body from infection anddisease. Examples of leukocytes include, without limitation,neutrophils, eosinophils, basophils, lymphocytes, and monocytes.

The term “mammal” includes, without limitation, humans, domestic animals(e.g., dogs or cats), farm animals (cows, horses, or pigs), monkeys,rabbits, mice, and laboratory animals.

The terms “modulate”, “modulation” and the like refer to the ability ofa compound to increase or decrease the function and/or expression of sykand/or JAK, where such function may include transcription regulatoryactivity and/or protein-binding. Modulation may occur in vitro or invivo. Modulation, as described herein, includes the inhibition,antagonism, partial antagonism, activation, agonism or partial agonismof a function or characteristic associated with syk and/or JAK, eitherdirectly or indirectly, and/or the upregulation or downregulation of theexpression of syk and/or JAK, either directly or indirectly. In apreferred embodiment, the modulation is direct Inhibitors or antagonistsare compounds that, e.g., bind to, partially or totally blockstimulation, decrease, prevent, inhibit, delay activation, inactivate,desensitize, or downregulate signal transduction. Activators or agonistsare compounds that, e.g., bind to, stimulate, increase, open, activate,facilitate, enhance activation, activate, sensitize or upregulate signaltransduction. The ability of a compound to inhibit the function of sykand/or JAK can be demonstrated in a biochemical assay, e.g., bindingassay, or a cell-based assay, e.g., a transient transfection assay.

“Modulators” of activity are used to refer to “ligands”, “antagonists”and “agonists” identified using in vitro and in vivo assays for activityand their homologs and mimetics. Modulators include naturally occurringand synthetic ligands, antagonists, agonists, molecules and the like.Assays to identify antagonists and agonists include, e.g., applyingputative modulator compounds to cells, in the presence or absence of areceptor of the invention and then determining the functional effects ona receptor of the invention activity. Samples or assays comprising areceptor of the invention that are treated with a potential activator,inhibitor, or modulator are compared to control samples without theinhibitor, activator, or modulator to examine the extent of effect.Control samples (untreated with modulators) are assigned a relativeactivity value of 100% Inhibition is achieved when the activity value ofa receptor of the invention relative to the control is about 80%,optionally 50% or 25-1%. Activation is achieved when the activity valueof a receptor of the invention relative to the control is 110%,optionally 150%, optionally 200-500%, or 1000-3000% higher.

“Patient” refers to human and non-human animals, especially mammals.Examples of patients include, but are not limited to, humans, cows,dogs, cats, goats, sheep, pigs and rabbits.

Turning next to the compositions of the invention, the term“pharmaceutically acceptable carrier or excipient” means a carrier orexcipient that is useful in preparing a pharmaceutical composition thatis generally safe, non-toxic and neither biologically nor otherwiseundesirable, and includes a carrier or excipient that is acceptable forveterinary use as well as human pharmaceutical use. A “pharmaceuticallyacceptable carrier or excipient” as used in the specification and claimsincludes both one and more than one such carrier or excipient.

The terms “pharmaceutically effective amount”, “therapeuticallyeffective amount” or “therapeutically effective dose” refers to theamount of the subject compound that will elicit the biological ormedical response of a tissue, system, animal or human that is beingsought by the researcher, veterinarian, medical doctor or otherclinician. The term “therapeutically effective amount” includes thatamount of a compound that, when administered, is sufficient to preventdevelopment of, or alleviate to some extent, one or more of the symptomsof the condition or disorder being treated. The therapeuticallyeffective amount will vary depending on the compound, the disorder orcondition and its severity and the age, weight, etc., of the mammal tobe treated.

The term “platelet” refers to a minute, normucleated, disklike cellfound in the blood plasma of mammals that functions to promote bloodclotting.

The terms “prevent”, “preventing”, “prevention” and grammaticalvariations thereof as used herein, refers to a method of partially orcompletely delaying or precluding the onset or recurrence of a disorderor condition and/or one or more of its attendant symptoms or barring asubject from acquiring or reacquiring a disorder or condition orreducing a subject's risk of acquiring or reacquiring a disorder orcondition or one or more of its attendant symptoms.

The term “recanalization” refers to the process of restoring flow to orreuniting an interrupted channel of the body, such as a blood vessel.

The term “restenosis” refers to a re-narrowing or blockage of an arteryat the same site where treatment, such as an angioplasty or a stentprocedure, has been performed.

The phrase “selectively” or “specifically” when referring to binding toa receptor, refers to a binding reaction that is determinative of thepresence of the receptor, often in a heterogeneous population ofreceptors and other biologics. Thus, under designated conditions, thecompounds bind to a particular receptor at least two times thebackground and more typically more than 10 to 100 times background.Specific binding of a compound under such conditions requires a compoundthat is selected for its specificity for a particular receptor. Forexample, small organic molecules can be screened to obtain only thosecompounds that specifically or selectively bind to a selected receptorand not with other receptors or proteins. A variety of assay formats maybe used to select compounds that are selective for a particularreceptor. For example, High-throughput screening assays are routinelyused to select compounds that are selective for a particular a receptor.

As used herein, the term “Sickle cell anemia” refers to an inheriteddisorder of the red blood cells in which both hemoglobin alleles encodethe sickle hemoglobin (S) protein, i.e., the S/S genotype. The presenceof abnormal hemoglobin results in the production of unusually shapedcells, which do not survive the usual length of time in the bloodcirculation. Thus, anemia results. “Anemia” refers to a decrease in thenumber of red blood cells and/or hemoglobin in the blood.

The term “Sickle cell disease” refers to an inherited disorder of thered blood cells in which one hemoglobin allele encodes the sicklehemoglobin (S) protein, and the other allele encodes another unusualhemoglobin protein, such as hemoglobin (S), (C), (D), (E), and (βThal).Examples of sickle cell disease genotypes include, without limitation,the S/S, S/C, S/D, S/E, and S/βThal genotypes. The most common types ofsickle cell disease include sickle cell anemia, sickle-hemoglobin Cdisease, sickle beta-plus thalassemia, and sickle beta-zero thalassemia.

The “subject” is defined herein to include animals such as mammals,including, but not limited to, primates (e.g., humans), cows, sheep,goats, horses, dogs, cats, rabbits, rats, mice and the like. Inpreferred embodiments, the subject is a human.

As used herein, the term “syk” refers to a spleen tyrosine kinase(RefSeq Accession No. P-043405) or a variant thereof that is capable ofmediating a cellular response to T-cell receptors in vitro or in vivo.syk variants include proteins substantially homologous to native syk,i.e., proteins having one or more naturally or non-naturally occurringamino acid deletions, insertions or substitutions (e.g., sykderivatives, homologs and fragments). The amino acid sequence of sykvariant preferably is at least about 80% identical to a native syk, morepreferably at least about 90% identical, and most preferably at leastabout 95% identical.

The term “syk inhibitor” refers to any agent that inhibits the catalyticactivity of spleen tyrosine kinase.

The term “thrombosis” refers to the blockage or clotting of a bloodvessel caused by a clumping of cells, resulting in the obstruction ofblood flow. The term “thrombosis” refers to the clot that is formedwithin the blood vessel.

The terms “treat”, “treating”, “treatment” and grammatical variationsthereof as used herein, includes partially or completely delaying,alleviating, mitigating or reducing the intensity of one or moreattendant symptoms of a disorder or condition and/or alleviating,mitigating or impeding one or more causes of a disorder or condition.Treatments according to the invention may be applied preventively,prophylactically, pallatively or remedially.

The term “vessel” refers to any channel for carrying a fluid, such as anartery or vein. For example, a “blood vessel” refers to any of thevessels through which blood circulates in the body. The lumen of a bloodvessel refers to the inner open space or cavity of the blood vessel.

2. EMBODIMENTS OF THE INVENTION a. Compounds

The present invention provides in one embodiment, a compound havingFormula (I):

or a tautomer or pharmaceutically acceptable salt thereof,wherein:Y^(1a) is selected from the group consisting of N, CH and C;Z^(1a) is selected from the group consisting of a bond, —N(C₁₋₄alkyl)-,—SO₂—, —CO—, —NR^(4d)SO₂—, heterocyclyl, heterocyclylcarbonyl andheterocyclylsulfonyl;R^(1a) is selected from the group consisting of:

(a) H;

(b) C₁₋₈alkyl, optionally substituted with from 1 to 3 substituentsselected from the group consisting of amino, hydroxy, C₁₋₈alkoxy,heterocyclyl, aminocarbonyl, aminoC₁₋₈alkoxy, aryl and heteroaryl;(c) C₃₋₈cycloalkyl, optionally substituted with from 1 to 3 aminosubstituents;(d) aryl, optionally substituted with from 1 to 3 substituents selectedfrom the group consisting of C₁₋₈alkyl, C₁₋₈alkoxy, C₁₋₈alkylamino,C₁₋₈alkylcarbonylamino; aminocarbonylC₁₋₈alkoxy, aminosulfonyl,aminocarbonyl, aminoC₁₋₈alkylene carbonylC₁₋₈alkoxy and halo;(e) heterocyclyl, halogens, cyano, optionally substituted with from 1 to3 substituents selected from the group consisting of C₁₋₈alkyl, oxo andC₁₋₈alkoxycarbonyl, cyano C₁₋₆alkylcarbonyl, aminocarbonyl,arylC₁₋₄alkoxycarbonyl, arylaminocarbonyl; and(f) heteroaryl, optionally substituted with from 1 to 3 substituentsselected from the group consisting of C₁₋₈alkyl, C₁₋₈alkylsulfonyl andcyanoC₁₋₈alkylenecarbonyl;R^(2a) is H, C₁₋₈alkyl, or is taken together with R^(1a) and thenitrogen to which each is attached to form a heterocyclic or heteroarylring, containing 1-3 heteroatoms, including N, O or S, optionallysubstituted with from 1 to 2 substituents, R^(2b), independentlyselected from the group consisting of C₁₋₈alkoxy, C₁₋₈alkoxycarbonyl,C₁₋₈alkoxycarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene,amino, aminoC₁₋₈alkylene, aminoaryl, aminocarbonyl, aminocarbonylamino,aminocarbonylC₁₋₈alkylene, aminocarbonylaminoC₁₋₈alkylene,aminocarboxyC₁₋₈alkylene, aminosulfonyl, aminosulfonylC₁₋₈alkylene,C₁₋₈alkyl, C₁₋₈alkylsulfonyl, C₁₋₈alkylsulfonylC₁₋₈alkylene,C₁₋₈alkylheterocyclyl, C₁₋₈alkylheterocyclylC₁₋₈alkylene, aryl,arylC₁₋₈alkoxycarbonylamino, carboxy, carboxyC₁₋₈alkylene, cyano,C₃₋₈cycloalkyl, halo, heteroaryl, heteroarylC₁₋₈alkylene, heterocyclyl,hydroxy, hydroxyC₁₋₈alkylene, hydroxycarbonylC₁₋₈alkylene, imino, oxoand ═S;R^(3a) is H, C₁₋₈alkyl, or is taken together with the moietyR^(4a)—Z^(1a) and the nitrogen to which each is attached to form aheterocyclic or heteroaryl ring, optionally substituted with from 1 to 2substituents R^(2b) and R^(2c) each of which is independently selectedfrom the group consisting of C₁₋₈alkoxy, C₁₋₈alkoxycarbonyl,C₁₋₈alkoxycarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene,amino, aminoC₁₋₈alkylene, aminoaryl, aminocarbonyl, aminocarbonylamino,aminocarbonylaminoC₁₋₈alkylene, aminocarboxyC₁₋₈alkylene, aminosulfonyl,aminosulfonylC₁₋₈alkylene, C₁₋₈alkyl, C₁₋₈alkylsulfonyl,C₁₋₈alkylsulfonylC₁₋₈alkylene, C₁₋₈alkylheterocyclyl,C₁₋₈alkylheterocyclylC₁₋₈alkylene, aryl, arylC₁₋₈alkoxycarbonylamino,carboxy, carboxyC₁₋₈alkylene, cyano, C₃₋₈cycloalkyl, halo, heteroaryl,heteroarylC₁₋₈alkylene, heterocyclyl, hydroxy, hydroxyC₁₋₈alkylene,imino, oxo and ═S;R^(4a) is selected from the group consisting of:(a) aryl, optionally substituted with from 1 to 3 substituents, R^(4c),each of which is independently selected from the group consisting ofC₁₋₈alkoxy, amino, C₁₋₈alkylcarbonyl and aminocarbonylC₁₋₈alkoxy;(b) heteroaryl, heterobicyclic C₁₋₈ alkyl, halo, hydroxyl, optionallysubstituted with from 1 to 3 substituents, R^(4c), each of which isindependently selected from the group consisting of C₁₋₈alkyl, halogen,hydroxyl, oxo C₁₋₈alkoxy and ═S;(c) heterocyclyl, each of which is optionally substituted with from 1 to3 substituents, R^(4c), each of which is independently selected from thegroup consisting of C₁₋₈alkyl and oxo;R^(4b) is selected from the group consisting of H, C₁₋₈alkyl,aminoC₁₋₈alkylene, C₁₋₈alkylcarbonyl, C₁₋₈alkylcarbonylamino,C₁₋₈alkylsulfonyl, C₁₋₈alkylsulfinyl, C₁₋₈alkylsulfonylamino,C₁₋₈alkylsulfonyl C₁₋₈alkylene, C₁₋₈alkylthio, C₁₋₈alkoxy,C₁₋₈alkoxyC₁₋₈alkyleneamino, C₁₋₈alkoxycarbonylamino,C₁₋₈alkoxycarbonyl, C₁₋₈alkoxyC₁₋₈alkylene,C₁₋₈alkoxycarbonylC₁₋₈alkylene, amino, aminocarbonyl,aminocarbonylC₁₋₈alkylene. aminosulfonyl,C₁₋₈alkoxycarbonylC₁₋₈alkyleneaminosulfonyl,aminoC₁₋₈alkyleneaminocarbonyl, aminocarbonylC₁₋₈alkoxy,aminoC₁₋₈alkylene, aminoC₁₋₈alkylenecarbonyl,aminocarbonylC₁₋₈alkyleneaminoesulfonyl, carboxy,carboxyC₁₋₈alkyleneaminosulfonyl, carboxyC₁₋₈alkylene,C₃₋₈cycloalkylcarbonylamino, C₃₋₈cycloalkylcarbonyl, halo, hydroxy,hydroxyC₁₋₈alkylene, aminoC₁₋₈alkyleneamino, aminoC₁₋₈alkylenecarbonyl,aminoC₁₋₈alkyleneaminocarbonylC₁₋₈alkylene,hydroxyC₁₋₈alkylenecarbonylamino, hydroxylC₁₋₈alkylenecarbonyl,hydroxyC₁₋₈alkyleneamino, hydroxyC₁₋₈alkyleneaminosulfonyl,hydroxyC₁₋₈alkyleneaminocarbonyl, oxo and heterocyclyl;if R^(4b) is heterocyclyl, it is optionally substituted with 1-3substituents, R^(4d), independently selected from the group consistingof C₁₋₈alkyl, C₁₋₈alkoxy, hydroxy, amino, halo, cyano, oxo, ═S,C₁₋₈alkoxycarbonyl, carboxy, aminocarbonyl, aminosulfonyl,C₁₋₈alkylcarbonyl and C₁₋₈alkylaminocarbonyl;R^(5a) is selected from the group consisting of H, C₁₋₈alkyl,C₃₋₈cycloalkyl and C₁₋₈alkylarylsulfonyl;R^(6a) is selected from the group consisting of H, C₁₋₈alkyl, halo,hydroxyl and oxo;R^(7a) is selected from the group consisting of H, C₁₋₈alkyl, cyano,C₁₋₈alkoxyC₁₋₈alkylene, C₂₋₈alkynyl, C₃₋₈cycloalkyl, C₁₋₈alkylcarbonyl,hydroxy, oxo, halo and aryl, wherein each aryl and heteroaryl can beoptionally substituted with halo, C₁₋₈alkyl, C₁₋₈alkoxy, cyano, amino,hydroxyl, heteroaryl; and the dashed line indicates a double or singlebond.

In another group of embodiments, R^(1a) is H. In another group ofembodiments, R^(1a) is C₁₋₈alkyl. In another group of embodiments,R^(1a) is C₃₋₈cycloalkyl. In another group of embodiments, R^(1a) iscyclohexyl, cyclopropyl or cyclobutyl. In another group of embodiments,R^(1a) is aryl. In another group of embodiments, R^(1a) is phenyl. Inanother group of embodiments, R^(1a) is heterocyclyl. In another groupof embodiments, R^(1a) is heteroaryl.

In another group of embodiments, R^(2a) is H. In another group ofembodiments, R^(2a) is C₁₋₈alkyl. In another group of embodiments,R^(2a) is taken together with R^(1a) and the nitrogen to which each isattached to form a heterocyclic ring selected from the group consistingof piperidine, piperazine, homopiperazine and pyrrolidine.

In another group of embodiments, each R^(2b) is independently selectedfrom the group consisting of —CH₂NH₂, —CH₂NHCH₃, —CH₂N(CH₃)₂,—CH₂CH₂NH₂, —C(CH₃)₂NH₂, —NH₂, —OH, —CH₂OH, —CH₂CH₂OH, —CH₂NHC(O)NH₂,—C(O)NH₂ and —CH₂C(O)NH₂, —CH(NH₂)CO₂CH₃, —CHNH₂)CH₂CO₂Et.

In another group of embodiments, Y¹ is N. In another group ofembodiments, Y^(1a) is CH. In another group of embodiments, Y^(1a) is C.

In another group of embodiments, R^(1a) is H. In another group ofembodiments, R^(1a) is C₁₋₈alkyl. In another group of embodiments,R^(1a) is taken together with the moiety R^(4a)—Z^(1a) and the nitrogento which each is attached to form a heterocyclic ring. In another groupof embodiments, R^(1a) is taken together with the moiety R^(4a)—Z^(1a)and the nitrogen to which each is attached to form a heteroaryl ring.

In another group of embodiments, R^(4a) is aryl. In another group ofembodiments, R^(4a) is phenyl. In another group of embodiments, R^(4a)is heteroaryl. In another group of embodiments, R^(4a) is heterocyclyl.

In another group of embodiments, Z¹ is a bond. In another group ofembodiments, Z¹ is —N(C₁₋₈alkyl)-. In another group of embodiments, Z¹is —SO₂—. In another group of embodiments, Z¹ is —CO—. In another groupof embodiments, Z¹ is —NR^(4d)SO₂—. In another group of embodiments, Z¹is heterocyclyl. In another group of embodiments, Z¹ isheterocyclylcarbonyl. In another group of embodiments, Z¹ isheterocyclylsulfonyl.

In another group of embodiments, R^(5a) is H.

In another group of embodiments, R^(6a) is H.

In another group of embodiments, R^(7a) is selected from the groupconsisting of H, cyano, phenyl and pyridyl.

In another group of embodiments, R^(4b) is selected from the groupconsisting of H, C₁₋₈alkyl, C₁₋₈alkylcarbonyl, C₁₋₈alkylcarbonylamino,C₁₋₈alkylsulfonyl, C₁₋₈alkylsulfinyl, C₁₋₈alkylsulfonylamino,C₁₋₈alkylsulfonyl C₁₋₈alkylene, C₁₋₈alkylthio, C₁₋₈alkoxy,C₁₋₈alkoxycarbonylamino, C₁₋₈alkoxycarbonyl, C₁₋₈alkoxycarbonylC₁₋₈alkylene, aminocarbonyl, aminosulfonyl, C₁₋₈alkoxycarbonylC₁₋₈alkyleneaminosulfonyl, aminoC₁₋₈alkyleneaminocarbonyl,aminoC₁₋₈alkyleneaminocarbonyl, aminocarbonylC₁₋₈alkoxy, aminocarbonyl,aminocarbonylC₁₋₈alkyleneaminosulfonyl, carboxy,carboxyC₁₋₈alkyleneaminosulfonyl, C₃₋₈cycloalkylcarbonylamino, halo,hydroxy, hydroxyC₁₋₈alkylene, hydroxyC₁₋₈alkylenecarbonylamino,hydroxyC₁₋₈alkyleneamino-, hydroxyC₁₋₈alkyleneaminosulfonyl,hydroxyC₁₋₈alkyleneaminocarbonyl, oxo and heterocyclyl.

The present invention provides in another embodiment, a compound, havingthe formula Ia:

or tautomer or a pharmaceutically acceptable salt thereof, wherein:

Y^(1b) is N, CH or C;

Z^(1b) is heterocyclyl, —NR^(4d)SO₂—, heterocyclylsulfonyl or a bond,R^(1b) is selected from the group consisting of

(a) H,

(b) C₁₋₈alkyl, optionally substituted with amino, aminoC₁₋₈alkyl,aminocarbonyl, alkoxy, C₁₋₈alkylaminocarbonyl, aminoC₁₋₈alkylene,aminoC₁₋₈alkoxy, aryl, hydroxy or heterocyclyl,(c) C₃₋₈cycloalkyl, optionally substituted with alkyl, amino, hydroxyl,C₁₋₈alkoxy, C₁₋₈alkylaminocarbonyl, aminoC₁₋₈alkylene, aminoC₁₋₈alkoxy;(d) aryl, optionally substituted with alkyl, amino, aminosulfonyl,aminoC₁₋₈alkylene, C₁₋₈alkylaminocarbonylC₁₋₈alkoxy, hydroxyl,C₁₋₈alkoxy, C₁₋₈alkylaminocarbonyl, aminoC₁₋₈alkylene, aminoC₁₋₈alkoxyor halo;(e) heterocyclyl, optionally substituted with C₁₋₈ alkyl,C₁₋₈alkoxycarbonyl, amino, hydroxyl, C₁₋₈alkoxy, C₁₋₈alkylaminocarbonyl,aminoC₁₋₈alkylene, aminoC₁₋₈alkoxy, oxo, or cyanoC₁₋₈alkylenecarbonyl;(f) heteroaryl, optionally substituted with alkyl, amino, hydroxyl,C₁₋₈alkoxy, C₁₋₈alkylaminocarbonyl, aminoC₁₋₈alkylene oraminoC₁₋₈alkoxy;R^(4b) is selected from the group consisting of H, hydroxy,C₁₋₈alkylcarbonyl-, C₁₋₈alkoxycarbonylC₁₋₈alkylene,C₁₋₈alkoxycarbonylamino, aminocarbonylC₁₋₈alkylene, hydroxycarbonyl,hydroxycarbonylC₁₋₈alkylene, aminocarbonyl, aminoC₁₋₈alkylene, hydroxy,hydroxyC₁₋₈alkylenecarbonylamino, C₁₋₈alkylaminoC₁₋₈alkylene,C₁₋₈alkylaminoC₁₋₈alkyleneaminocarbonyl, aminocarbonylamino,C₁₋₈alkylsulfonylC₁₋₈alkylene, C₃₋₈cycloalkylcarbonylamino andC₁₋₈alkylcarbonylamino; andR^(4c) is H, amino, C₁₋₈alkylcarbonyl, aminocarbonylC₁₋₈alkoxy;R^(4d) is H or C₁₋₈alkyl;R⁵ is H or C₁₋₈alkyl;R⁶ is selected from the group consisting of H, C₁₋₈alkyl, halo, hydroxyand oxo;R⁷ is selected from the group consisting of H, C₁₋₈alkyl, cyano,C₁₋₈alkoxyC₁₋₈alkylene, C₁₋₈alkylcarbonyl, hydroxy, oxo, halo, aryl andheteroaryl; and the dashed line indicates a double or single bond; orthe moiety R^(4b)—Z^(1b) combines with R^(4c) to form 5-7 membraneheterocyclic ring, containing 1-3 heteroatoms, including N, O, S,optionally substituted with C₁₋₈alkyl, halo, cyano, oxo, ═S, amino,hydroxyl, C₁₋₈alkylcarbonyl, C₁₋₈alkoxy or aryloxycarbonylC₁₋₈alkylene.

The present invention provides in another embodiment, a compound, havingthe formula selected from the group consisting of:

wherein R^(1b) is C₁₋₈alkyl, optionally substituted with amino,heterocyclyl or aminoheterocyclyl, and C₃₋₈cycloalkyl, optionallysubstituted with amino; R⁷ is halo, hydroxy, C₁₋₈alkoxyC₁₋₈alkylene,C₁₋₈alkylcarbonyl, cyano or phenyl, pyridinyl; and R⁶ is H, C₁₋₈alkyl orhalo; R⁹ is H or C₁₋₈alkyl; and R¹⁰ is H, C₁₋₈alkyl, C₁₋₈alkoxy,C₁₋₈alkoxycarbonylC₁₋₈alkylene, aminoC₁₋₈alkylene,aminocarbonylC₁₋₈alkylene, carboxyC₁₋₈alkylene, C₃₋₈cycloalkyl andhydroxyC₁₋₈alkylene; or a tautomer or a pharmaceutically acceptable saltthereof.

In another group of embodiments, R^(1b) is C₁₋₈alkyl, optionallysubstituted. In another group of embodiments, R^(1b) is C₃₋₈cycloalkyl,optionally substituted. In another group of embodiments, R^(1b) iscyclohexyl, cyclopropyl or cyclobutyl, each of which is optionallysubstituted. In another group of embodiments, R^(1b) is heterocyclyl,optionally substituted. In another group of embodiments, R^(1b) isheterocyclyl, which is selected from the group consisting ofpiperidinyl, morpholino and tetrahydrothiophenyl. In another group ofembodiments, R^(1b) is aryl. In another group of embodiments, R^(1b) ismonocyclic heteroaryl. In another group of embodiments, R^(1b) ispyrazole.

In another group of embodiments, Z^(1b) is heterocyclyl, which isselected from the group consisting of:

In another group of embodiments, the moiety

is selected from the group consisting of:

wherein R⁸ is selected from the group consisting of H, C₁₋₈alkyl,hydroxycarbonylC₁₋₈alkylene, alkoxycarbonylC₁₋₈alkylene andC₁₋₈alkylaminoC₁₋₈alkyleneaminocarbonylC₁₋₈alkylene.

The present invention provides in another group of embodiments, acompound having the formula:

wherein R^(2c) is aminocarbonyl, aminoC₁₋₈alkylene or hydroxy; and R⁷ isselected from the group consisting of halo, hydroxy,C₁₋₈alkoxyC₁₋₈alkylene, C₁₋₈alkylcarbonyl, cyano, phenyl and pyridinyl.

The present invention provides in another group of embodiments, acompound having the formula:

The present invention provides in another group of embodiments, acompound having the formula:

wherein R^(1b) is C₁₋₈alkyl, C₃₋₈cycloalkyl, aryl or heteroaryl; each ofwhich is optionally substituted with hydroxy, C₁₋₈alkoxy oraminoC₁₋₈alkoxy; R^(4b) is H or CH₃CO—; R⁷ is H, C₁₋₈alkyl,C₁₋₈alkylcarbonyl, aminocarbonyl, cyano, phenyl or pyridinyl; R⁹ is H orC₁₋₈alkyl; and R¹⁰ is H, C₁₋₈alkyl, C₁₋₈alkoxy,C₁₋₈alkoxycarbonylC₁₋₈alkylene, aminoC₁₋₈alkylene,aminocarbonylC₁₋₈alkylene, carboxyC₁₋₈alkylene, C₃₋₈cycloalkyl andhydroxyC₁₋₈alkylene.

The present invention provides in another group of embodiments, acompound having the formula Ib:

or a tautomer or pharmaceutically acceptable salt thereof wherein:

Y^(1b) is C or N;

each R^(2b) is independently selected from the group consisting ofamino, aminoC₁₋₈alkylene, aminocarbonyl, aminocarbonylC₁₋₈alkylene,carboxy, carboxyC₁₋₈alkylene, C₁₋₈alkoxycarbonyl,C₁₋₈alkoxycarbonylC₁₋₈alkylene and hydroxy;R^(4b) is C₁₋₈alkylcarbonyl;Z^(1b) is -heterocyclyl;

R^(4c) H;

R^(7b) is H, cyano, pyridinyl or null;m is 0 or 1; andp is 0 or 1.

The present invention provides in another group of embodiments, acompound having the formula Ic:

or a tautomer or a pharmaceutically acceptable salt thereof; wherein:R^(2b) is selected from the group consisting of H, aminoC₁₋₈alkylene,aminocarbonyl, aminocarbonylC₁₋₈alkylene, hydroxy,hydroxycarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonyl andC₁₋₈alkoxycarbonylC₁₋₈alkylene;

R^(4b) is H or CH₃CO—;

R^(7b) is H, cyano or pyridinyl; andm is 0 or 1.

The present invention provides in another group of embodiments, acompound having the formula Id:

or a tautomer or a pharmaceutically acceptable salt thereof; wherein:

Y^(1b) is C or N;

R^(2b) is selected from the group consisting of H, amino,aminoC₁₋₈alkylene, aminocarbonyl, aminocarbonylC₁₋₈alkylene, hydroxy,hydroxycarbonylC₁₋₈alkylene, carboxy, C₁₋₈carboxyalkylene,C₁₋₈alkoxycarbonyl and C₁₋₈alkoxycarbonylC₁₋₈alkylene;R^(7a) is H, C₁₋₈alkyl, halo, hydroxy, C₁₋₈alkoxyC₁₋₈alkylene,C₁₋₈alkylcarbonyl, cyano or pyridinyl; andm is 0 or 1.

In another group of embodiments, m is 0; and R^(2a) is selected from thegroup consisting of H, cyano or pyridinyl. In another group ofembodiments, m is 1; and R^(7a) is selected from the group consisting ofH, cyano or pyridinyl.

The present invention provides in another group of embodiments, acompound having the formula:

and R^(4b) is H or CH₃CO—.

The present invention provides in another group of embodiments, acompound having the formula Ie:

or a tautomer or pharmaceutically acceptable salt thereof, wherein:R^(2b) is selected from the group consisting of H, aminoC₁₋₈alkylene,aminocarbonyl, aminocarbonylC₁₋₈alkylene, hydroxy,hydroxycarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonyl andC₁₋₈alkoxycarbonylC₁₋₈alkylene;R^(7b) is H, cyano, or pyridinyl; andm is 0 or 1.

The present invention provides in another embodiment, a compound havingthe formula:

or a tautomer or a pharmaceutically acceptable salt thereof.

The present invention provides in another group of embodiments, acompound having the formula II:

or a tautomer or a pharmaceutically acceptable salt thereof, wherein:R^(1c) is selected from the group consisting of(a) C₃₋₈cycloalkyl, and(b) aryl, each of which is optionally substituted with from 1 to 2substituents selected from the group consisting of C₁₋₈alkyl, amino andhydroxyl; oris taken together with R^(2c) to form a heterocyclic ring, containing1-3 heteroatoms, including N, O, S, optionally substituted with from 1to 2 substituents, each of which is independently selected from thegroup consisting of aminocarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonyl,aminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene,C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene, amino,aminocarbonylaminocarbonylamino, arylC₁₋₈alkoxycarbonylamino,aminocarbonylamino and oxo;R^(2c) is H or is taken together with R^(1c) to form a heterocyclicring, optionally cyanoC₁₋₈alkylcarbonyl, cyanoC₁₋₆alkylcarbonylaminosubstituted with from 1 to 2 substituents independently selected fromthe group consisting of aminocarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonyl,aminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene,C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene, amino,aminocarbonylaminocarbonylamino, arylC₁₋₈alkoxycarbonylamino,aminocarbonylamino and oxo;

R^(3c) is H or taken together with R^(4d) to form a heterocyclic orheteroaryl ring, each of which is optionally substituted with from 1 to2 substituents independently selected from the group consisting ofC₁₋₈alkyl, C₁₋₈alkylheterocyclyl, aminoC₁₋₈alkylene, aminoaryl,hydroxyC₁₋₈alkylene, aminocarbonyl, C₁₋₈alkoxycarbonyl, amino, imino,C₁₋₈alkylcarbonylamino, oxo, halo, aryl, heterocyclyl,heterocyclylC₁₋₈alkylene and C₁₋₈alkylheterocyclylC₁₋₈alkylene;

R^(4d) is independently selected from the group consisting of(a) C₃₋₈cycloalkyl,(b) aryl,(c) heteroaryl, and(d) heterocyclyl; each of which is optionally substituted with from 1 to3 substituents R^(4e), R^(4f) and —Z^(1c)R^(4g), each of which isindependently selected from the group consisting of C₁₋₈alkyl,aminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene, aminocarbonyl,C₁₋₈alkoxycarbonyl, carboxy, amino, imino, C₁₋₈alkylcarbonylamino, oxo,halo, aryl, heterocyclyl and heterocyclylC₁₋₈alkylene, provided that atleast one of R^(1c) and R^(2c) or R^(3c) and R^(4d) are combined withthe nitrogen to which each is attached to form a heterocyclyl orheteroaryl ring; andR^(7c) is selected from the group consisting of H, C₁₋₈alkyl,C₂₋₈alkynyl, cyano, aminocarbonyl, nitro, C₁₋₈alkoxy, halogen, aryl,heteroaryl and C₃₋₈cycloalkyl.

The present invention provides in another group of embodiments, acompound having the formula IIa:

or a tautomer or a pharmaceutically acceptable salt thereof.

In another group of embodiments, R^(3c) is H.

In another group of embodiments, R^(4d) is phenyl. In another group ofembodiments, R^(4d) is indazyl. In another group of embodiments, R^(4d)is cyclohexyl. In another group of embodiments, R^(4d) is dihydroindoyl.

In another group of embodiments, R^(3c) s taken together with R^(4d) andthe nitrogen to which is each is attached to form heterocyclic ringselected from the group consisting of pyridinyl, imidazolyl,tetrahydroimidazoyl, indazolyl, piperidinyl, piperazinyl, pyrrolidinyl,triazoyl and benzamidazoyl.

In another group of embodiments, R^(4d)R³N— is selected from the groupconsisting of:

the dashed lined indicates a single or double bond; and the wavy lineindicates the point of attachment to the remainder of the molecule.

The present invention provides in another group of embodiments, acompound having the formula IIb:

or a tautomer or a pharmaceutically acceptable salt thereof, wherein:R^(7c) is halo;R^(1c) and R^(2c) are taken together to form a heterocyclic ring,optionally substituted with from 1 to 2 substituents independentlyselected from the group consisting of: aminocarbonylC₁₋₈alkylene,C₁₋₈alkoxycarbonyl, aminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene,C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene, amino,aminocarbonylaminocarbonylamino, arylC₁₋₈alkoxycarbonylamino,aminocarbonylamino and oxo;Z^(1c) is heterocyclyl, —N(C₁₋₄alkyl)-, —SO₂— or —CO—;R^(4g) is selected from the group consisting of H, C₁₋₈alkoxy,C₁₋₈alkylcarbonyl, C₁₋₈alkylcarbonylamino, amino,aminocarbonylaminoC₁₋₈alkylene, aminocarbonyl and aminosulfonyl;each R^(4e) and R^(4f) is independently selected from the groupconsisting of H—, C₁₋₈alkoxy and C₁₋₈alkylcarbonyl; or can be takentogether with —Z^(1c)—R^(4g) and the benzene ring to which each isattached to form a fused heterocyclic ring system, optionallysubstituted with from 1 to 2 oxo substituents, halogens, C₁₋₈alkyl,C₁₋₈alkyl.

The present invention provides in another group of embodiments, acompound, wherein the moiety

is selected from the group consisting of:

In another group of embodiments, R^(7c) is selected from the groupconsisting of F, Cl, Br, cyano and aminocarbonyl. In another group ofembodiments, R^(7c) is CONH₂ or F. In another group of embodiments,R^(1c) is C₃₋₈cycloalkyl. In another group of embodiments, R^(1c) iscyclohexyl. In another group of embodiments, R^(1c) is cyclopropyl. Inanother group of embodiments, R^(1c) is cyclobutyl. In another group ofembodiments, R^(1c) is aryl. In another group of embodiments, R^(1c) isphenyl.

In another group of embodiments, R^(2c) is taken together with R^(1c)and the nitrogen to which each is attached to form a pyrrolidinyl,piperidinyl or piperazinyl, diazapenyl ring.

The present invention provides in another group of embodiments, acompound having the formula:

wherein Y^(2b) is N, NH, C or CH; R^(2d) is H,C₁₋₈alkylaminoC₁₋₈alkylene, aminocarbonyl, aminocarbonylamino,aminocarbonylaminocarbonylamino, amino, oxo, aminoC₁₋₈alkylene,aminocarbonylaminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene,arylalkoxycarbonylamino; cyanoC₁₋₈alkylenecarbonyl,cyanoC₁₋₈alkylenecarbonylamino, hydroxyl, C₁₋₈alkoxycarbonyl,alkoxycarbonylC₁₋₈alkylene, optionally substituted by amino; and R^(7c)is H, halo, aminocarbonyl, cycloalkyl, cyano or pyridinyl; R⁹ is H orC₁₋₈alkyl; R¹⁰ is H, C₁₋₈alkyl, C₁₋₈alkoxy,C₁₋₈alkoxycarbonylC₁₋₈alkylene, aminoC₁₋₈alkylene,aminocarbonylC₁₋₈alkylene, carboxyC₁₋₈alkylene, C₃₋₈cycloalkyl andhydroxyC₁₋₈alkylene; m is 0 or 1; and q is 0 or 1.

In another group of embodiments, R^(7c) is selected from the groupconsisting of F, Cl, Br, cyano and aminocarbonyl. In another group ofembodiments, R^(7c) is CONH₂ or F.

The present invention provides in another group of embodiments, acompound having the formula:

or a tautomer or a pharmaceutically acceptable salt thereof.

The present invention provides in another embodiment, a compoundselected from the group consisting ofN2-(1H-indazol-6-yl)-N4-methyl-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;1-(4-(4-(4-(methylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N2-(1H-indazol-6-yl)-N4-methyl-5-(pyridin-3-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;1-(4-(4-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;4-(4-aminophenyl)-1-(5-carbamoyl-4-(cyclopropylamino)pyrimidin-2-yl)pyridinium;1-(4-aminophenyl)-3-(5-carbamoyl-4-(cyclopropylamino)pyrimidin-2-yl)-1H-imidazol-3-ium;2-(6-amino-7-chloro-1H-indazol-1-yl)-4-(cyclopropylamino)pyrimidine-5-carboxamide;1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide;1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide;4-(4-(aminomethyl)piperidin-1-yl)-N-(4-(piperazin-1-yl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine;1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;(S)-1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)pyrrolidin-3-ol;(S)-1-(4-(4-(4-(3-hydroxypyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;1-(4-(4-(6-(4-(aminomethyl)piperidin-1-yl)-9H-purin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-yl)-1H-indazol-6-amine;4-(4-(aminomethyl)piperidin-1-yl)-5-fluoro-N-(3,4,5-trimethoxyphenyl)pyrimidin-2-amine;1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N-(4-(piperazin-1-yl)phenyl)-4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amineand1-(4-(4-(4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N-(4-(piperazin-1-yl)phenyl)-4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amineand1-(4-(4-(4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamideand1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)pyrimidine-5-carboxamide;1-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidine-3-carboxamide;1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidine-3-carboxamide;tert-butyl(1-(5-fluoro-2-(4-(N-methylacetamido)phenylamino)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate;N-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)-N-methylacetamide;tert-butyl(1-(2-(4-carbamoylphenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)methylcarbamate;4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzamide;4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzenesulfonamide;6-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;1-(4-(4-(4-(2-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;tert-butyl(1-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidin-2-yl)methylcarbamateandN-(4-(2-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-yl)-1H-indazol-6-amine;1-((1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)methyl)urea;1-(4-(4-(5-fluoro-4-(4-(hydroxymethyl)piperidin-1-yl)pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;6-(5-fluoro-4-(4-(hydroxymethyl)piperidin-1-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;2-(1H-indazol-6-ylamino)-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;5-(1-butoxyethyl)-N4-cyclopropyl-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;1-(2-(1H-indazol-6-ylamino)-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-5-yl)ethanone;N4-cyclopropyl-N2-(1H-indazol-6-yl)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;1-(4-(4-(4-(piperidin-4-ylmethylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(piperidin-4-ylmethylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;N-(4-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;6-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;5-bromo-N4-cyclobutyl-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;5-chloro-N4-cyclobutyl-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;2-(1H-indazol-6-ylamino)-4-(cyclobutylamino)-6,7-dihydropyrrolo[2,3-d]pyrimidin-5-one;1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide;1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide;4-(4-(aminomethyl)piperidin-1-yl)-N-(4-(piperazin-1-yl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine;1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;(S)-4-(2-(aminomethyl)pyrrolidin-1-yl)-N-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine;(S)-1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)pyrrolidin-3-ol;(S)-1-(4-(4-(4-(3-hydroxypyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(3-(hydroxymethyl)piperidin-1-yl)pyrimidine-5-carboxamide;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(2-(2-hydroxyethyl)piperidin-1-yl)pyrimidine-5-carboxamide;1-(4-(4-(6-(4-(aminomethyl)piperidin-1-yl)-9H-purin-2-ylamino)phenyl)piperazin-1-yl)ethanone;(S)-4-(3-aminopiperidin-1-yl)-N-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine;(S)-1-(4-(4-(4-(3-aminopiperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-yl)-1H-indazol-6-amine;4-(4-(aminomethyl)piperidin-1-yl)-5-fluoro-N-(3,4,5-trimethoxyphenyl)pyrimidin-2-amine;1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N-(4-(piperazin-1-yl)phenyl)-4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine;1-(4-(4-(4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N-(4-(piperazin-1-yl)phenyl)-4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine;1-(4-(4-(4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide;1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)pyrimidine-5-carboxamide;1-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidine-3-carboxamide;1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidine-3-carboxamide;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(2-hydroxyethyl)-1,4-diazepan-1-yl)pyrimidine-5-carboxamide;tert-butyl(1-(5-fluoro-2-(4-(N-methylacetamido)phenylamino)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate;tert-butyl(1-(2-(4-carbamoylphenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)methylcarbamate;4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzamide;4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzenesulfonamide;N-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)-N-methylacetamide;6-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;1-(4-(4-(4-(2-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;tert-butyl(1-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidin-2-yl)methylcarbamate;andN-(4-(2-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-yl)-1H-indazol-6-amine.

The present invention provides in another embodiment, a compoundselected from the group consisting of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol;butyl2-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-N-methylphenylsulfonamido)acetate;2-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonamido)acetamide;2-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-N-methylphenylsulfonamido)aceticacid;1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperidin-4-ol;tert-butyl4-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)piperidine-1-carboxylate;N2-(1H-indazol-6-yl)-N4-(piperidin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;N4-cyclobutyl-N2-(1H-indazol-6-yl)-6-methyl-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;N4-(3-amino-2,2-dimethylpropyl)-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;N2-(1H-indazol-6-yl)-N4-(3-morpholinopropyl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;N-(4-(4-(4-aminobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;N-(4-(4-(2-aminocyclohexylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;N-(4-(4-(2-aminoethylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;N-(4-(4-(5-aminopentylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;1-(4-(4-(4-(4-aminocyclohexylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N-(4-(4-(5-aminopentylamino)-5-bromo-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;N-(4-(4-(5-aminopentylamino)-5,6-dibromo-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;N-(4-(4-(3-aminosulfonyl-phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;Ethyl2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate;2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)aceticacid;2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)-N-(2-(dimethylamino)ethyl)acetamide;methyl4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl(methyl)carbamate;methyl4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl(methyl)carbamate;N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-hydroxy-N-methylacetamide;N-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-hydroxy-N-methylacetamide;N⁴-cyclobutyl-N²-(4-methylsulfonylmethyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine;N⁴-cyclobutyl-N²-(3-methylsulfonylmethyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine;2-(3-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)phenoxy-N-methylacetamide;N²-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine;N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide;N-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide;N-(4-(4-(methylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide;2-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)acetamide;N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-(dimethlamino)-N-methylacetamide;N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-N-(3-dimethylaminopropyl)benzamide;N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-3-hydroxy-N-methypropanamide;N-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-3-hydroxy-N-methypropanamide;N²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;N²-(1H-indazol-6-ylamino)-N⁴-(2-methoxyethyl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;1-(4-(4-(4-(2-methoxyethylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N²-(1H-indazol-6-ylamino)-N⁴-(2-hydroxyethyl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;N⁴-(2-2-(aminoethoxy)ethyl)-N²-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;1-(4-(4-(4-(2-(2-aminoethoxy)amino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;(S)-1-(4-(4-(2-(2-aminoethoxy)ethylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)pyrrolidine-2-carboxamide;3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide;3-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide;3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamide;3-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamide;N-(4-(4-(3-(aminomethyl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;(R)-1-(4-(4-(4-(1-hydroxypropan-2-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;(R)-2-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol;(S)-1-(4-(4-(4-(1-hydroxypropan-2-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;(S)-2-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol;1-(4-(4-(4-(1,1-dioxy-tetrahydrothiophen-3-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;N2-(4-(piperazin-1-yl)phenyl)-N4-(1,1-dioxy-tetrahydrothiophen-3-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine;4-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide;4-(4-amino-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzoicacid;4-(cyclopropylamino)-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;4-(5-cyano-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide;N-(4-(5-cyano-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide;4-(5-cyano-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzenesulfonamide;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;4-(cyclopropylamino)-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide;4-(4-(1-(2-cyanoacetyl)piperidin-4-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide;4-(4-(1-(2-cyanoacetyl)piperidin-3-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide;4-(4-methoxyphenylamino)-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;6-(4-(4-fluorophenylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-1-methyl-3,4-dihydroquinolin-2-(1H)-one;6-(4-Amino-5-(4-fluorophenyl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-1-methyl-3,4-dihydroquinolin-2(1H)-one;6-(4-(benzylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-1-methyl-3,4-dihydroquinolin-2(1H)-one;4-(Benzylamino)-2-(1-methyl-2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile;Butyl1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylate;1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylicacid;1-(2-(4-(piperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylicacid;1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxamide;butyl2-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)acetate;2-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)aceticacid;2-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)acetamide;1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-chloropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;6-(5-fluoro-4-(3-oxopiperazin-1-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;1-(4-(4-(5-fluoro-4-(piperazin-1-yl)pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;4-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperazine-1-carboxamide;2-(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)acetamide;Benzyl1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-ylcarbamate;1-(4-(4-(4-(4-aminopiperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;1-(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)urea;2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(ureidomethyl)piperidin-1-yl)pyrimidine-5-carboxamide;4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzoicacid;6-(4-(4-(aminomethyl)piperidin-1-yl)-5-bromopyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;4-(4-(4-((dimethylamino)methyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzamide;ethyl3-amino-3-(1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)propanoate;methyl2-amino-2-(1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)acetate;N-(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)-2-cyanoacetamide;3-(4-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperazin-1-yl)-3-oxopropanenitrile;6-(4-(4-(aminomethyl)piperidin-1-yl)-5-(pyridin-4-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;6-(4-(4-(aminomethyl)piperidin-1-yl)-5-cyclopropylpyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;6-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one;4-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)benzamide;1-(4-(4-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone;6-(4-((1s,4s)-4-aminocyclohexylamino)-5-(pyridin-4-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-oneand6-(4-((1s,4s)-4-aminocyclohexylamino)-5-cyclopropylpyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one.

The present invention provides in another embodiment, a compound of theexamples.

The present invention provides in another embodiment, a compound of anyone of the tables.

The present invention provides in another embodiment, a compound of anyone of the figures.

It is understood that in another group of embodiments, any of the aboveembodiments may also be combined with other embodiments listed herein,to form other embodiments of the invention.

b. Methods of Synthesis

The compounds of the present invention may be prepared by known organicsynthesis techniques, including the methods described in more detail inthe Examples. In general, the compounds of structure (I) above may bemade by the following FIG. 1, wherein all substituents are as definedabove unless indicated otherwise.

Compounds having formula I may be prepared according to FIG. 1.2,4-dichloropyrrolopyrimidine 1.1 is protected with a protecting group,such as a tosyl group. Selective displacement of the 4-chloro group ofthe 2,4-dichloropyrrolopyrimidine by an appropriate amine, such asR²—NH—R¹ (available commercially or synthesized using methods known tothose skilled in the art), under basic conditions, such as withdiisopropylamine (DIA), provides compounds of formula 1.3. Subsequentdisplacement of the chloro group with an appropriate amine, such asR⁴—NH_R³ (available commercially or synthesized using methods known tothose skilled in the art), gives compound 1.4. Subsequent hydrolysis ofthe protecting group with hydroxide in an alcohol solvent gives thedesired product I, wherein R¹, R², R³ and R⁴ are as previously defined,wherein R¹, R², R³ and R⁴ are as previously defined.

Compounds having formula II may be prepared according to FIG. 2.Carboxylic acid 2.1 is converted to acid chloride 2.2 via a one-stepprocedure by treatment with a chlorination agent, such as thionylchloride, and esterification with an alcohol, such as ethanol, to formcompound 2.3 using conditions similar to that described below. Ester 2.3is dichlorinated with a chlorinating agent, such as phosphorousoxychloride. Selective displacement of the 4-chloro group of the2,4-dichloropyrimidine by an appropriate amine, such as R²—NH—R¹(available commercially or synthesized using methods known to thoseskilled in the art), under basic conditions, such as withdiisopropylamine (DIA), provides compounds of formula 2.5. Subsequenthydrolysis of the ester, displacement of the second chloro group withEDC and treatment with ammonia gives compound 2.7. Benzotriazolyl ethercompound 2.7 may also be prepared through a linear route. Displacementof the benzotriazolyl ether group with an appropriate amine, such asR⁴—NH_R³ (available commercially or synthesized using methods known tothose skilled in the art), gives the desired product IIa, wherein R¹,R², R³ and R⁴ are as previously defined.

Compounds having formula II may be prepared according to FIG. 3.Selective displacement of the 4-chloro group of the2,4-dichloropyrimidine by an appropriate amine, such as R²—NH—R¹(available commercially or synthesized using methods known to thoseskilled in the art), under basic conditions, such as withdiisopropylamine (DIA), provides compounds of formula 3.2. Displacementof the chloro group with an appropriate amine, such as R⁴—NH—R³(available commercially or synthesized using methods known to thoseskilled in the art), gives the desired product IIb, wherein R¹, R², R³and R⁴ are as previously defined.

One skilled in the art will recognize that in certain embodiments ofstructures (I-II) when R¹ or R² comprises a terminal heteroatom, it maybe advantageous to use a protecting group strategy. The protecting groupcan be removed using methods known to those skilled in the art to yieldcompounds of structure (I-II).

The compounds of the present invention may generally be utilized as thefree base. Alternatively, the compounds of this invention may be used inthe form of acid addition salts as described below.

c. Inhibition of syk and JAK Kinases

The activity of a specified compound as an inhibitor of a JAK kinase maybe assessed in vitro or in vivo. In some embodiments, the activity of aspecified compound can be tested in a cellular assay. Selectivity couldalso be ascertained in biochemical assays with isolated kinases.

Similar types of assays can be used to assess JAK kinase inhibitoryactivity and to determine the degree of selectivity of the particularcompound as compared to syk kinase. One means of assaying for suchinhibition is detection of the effect of the compounds of the presentinvention on the upregulation of downstream gene products. In theRamos/IL4 assay, B-cells are stimulated with the cytokine Interleukin-4(IL-4) leading to the activation of the JAK/Stat pathway throughphosphorylation of the JAK family kinases, JAK1 and JAK3, which in turnphosphorylate and activate the transcription factor Stat-6. One of thegenes upregulated by activated Stat-6 is the low affinity IgE receptor,CD23. To study the effect of inhibitors (e.g., the 2,4-substitutedpyrimindinediamine compounds described herein) on the JAK1 and JAK3kinases, human Ramos B-cells are stimulated with human IL-4. 10′post-stimulation, cells are subjected to intracellular flow cytometry tomeasure the extent of STAT-6 phosphorylation. 20 to 24 hourspost-stimulation, cells are stained for upregulation of CD23 andanalyzed using flow cytometry. A reduction of the amount ofphosphohorylated STAT-6 and/or cell surface CD23 present compared tocontrol conditions indicates that the test compound actively inhibitsthe JAK kinase pathway.

Additionally, IL-6 stimulation of Ramos B-cells induces JAKs 1, 2, andTyk2, leading to Stat-3 and Erk phosphorylation. 10′ post-stimulation,cells are subjected to intracellular flow cytometry to measure theability of compound to inhibit these phosphorylation events. Tospecifically measure the activity of JAK2, the CellSensor irfl-bla HELcell line expressing the beta-lactamase reporter gene controlled byStat5 will be used (Invitrogen, Carlsbad, Calif.). These cells express aconstitutively active JAK2 mutant (JAK2V617F), found naturally inmyeloproliferative neoplasms (Constantinescu, S., et. al, Trends BiochemSci., 2008; 33:122-31). A reduction in the amount of beta-lactamasereporter gene expression is used a measure of the JAK2 inhibitoryactivity of compounds.

The activity of the compounds of the invention may additionally becharacterized by assaying the effect of the compounds of the presentinvention described herein on A549 lung epithelial cells and U937 cells.A549 lung epithelial cells and U937 cells up-regulate ICAM-1 (CD54)surface expression in response to a variety of different stimuli.Therefore, using ICAM-1 expression as readout, test compound effects ondifferent signaling pathways can be assessed in the same cell type.Stimulation with IL-1β through the IL-1βreceptor activates theTRAF6/NFκB pathway resulting in up-regulation of ICAM-1. IFN.gammainduces ICAM-1 up-regulation through activation of the JAK1/JAK2pathway. The up-regulation of ICAM-1 can be quantified by flow cytometryacross a compound dose curve and EC₅₀ values are calculated.

The activity of the compounds of the invention may additionally becharacterized by assaying the effect of the compounds of the presentinvention described herein on A549 lung epithelial cells and U937 cells.A549 lung epithelial cells and U937 cells up-regulate ICAM-1 (CD54)surface expression in response to a variety of different stimuli.Therefore, using ICAM-1 expression as readout, test compound effects ondifferent signaling pathways can be assessed in the same cell type.Stimulation with IL-1β through the IL-1βreceptor activates theTRAF6/NFκB pathway resulting in up-regulation of ICAM-1. IFN.gammainduces ICAM-1 up-regulation through activation of the JAK1/JAK2pathway. The up-regulation of ICAM-1 can be quantified by flow cytometryacross a compound dose curve and EC₅₀ values are calculated. Exemplaryassays of this type are described in greater detail in the Examples.

Active compounds as described herein generally inhibit the JAK kinasepathway with an IC₅₀ in the range of about 1 mM or less, as measured inthe assays described herein. Of course, skilled artisans will appreciatethat compounds which exhibit lower IC₅₀s, (on the order, for example, of100 μM, 75 μM, 50 μM, 40 μM, 30 μM, 20 μM, 15 μM, 10 μM, 5 μM, 1 μM, 500nM, 100 nM, 10 nM, 1 nM, or even lower) can be particularly useful intherapeutic applications. In instances where activity specific to aparticular cell type is desired, the compound can be assayed foractivity with the desired cell type and counter-screened for a lack ofactivity against other cell types. The desired degree of “inactivity” insuch counter screens, or the desired ratio of activity vs. inactivity,may vary for different situations and can be selected by the user.

The active compounds also typically inhibit IL-4 stimulated expressionof CD23 in B-cells with an IC₅₀ in the range of about 2011M or less,typically in the range of about 10 μM, 1 μM, 500 nM, 100 nM, 10 nM, 1nM, or even lower. A suitable assay that can be used is the assaydescribed in the Examples, “Assay for Ramos B-cell Line Stimulated withIL-4.”In certain embodiments, the active compounds of the presentinvention have an IC₅₀ of less than or equal to 5 μM, greater than 5 μMbut less than 20 μM, greater than 20 μM, or greater than 20 μM but lessthan 50 μM in the assay described in the Examples.

The active compounds also typically inhibit expression of ICAM1 (CD54)induced by IFN.gamma exposure in U937 or A549 cells with an IC₅₀ in therange of about 20 μM or less, typically in the range of about 10 μM, 1μM, 500 nM, 100 nM, 10 nM, 1 nM, or even lower. The IC₅₀ againstexpression of ICAM (CD54) in IFN.gamma stimulated cells can bedetermined in a functional cellular assay with an isolated A549 or U937cell line. Suitable assays that can be used are the assays described inthe Examples, “A549 Epithelial Line Stimulated with IFNγ” and “U937IFN.gamma ICAM1 FACS Assay,” respectively. In certain embodiments, theactive compounds of the present invention have an IC₅₀ of less than orequal to 20 μM, greater than 20 μM, or greater than 20 μM but less than50 μM in the assays described in the Examples.

d. Compositions and Methods of Administration

The present invention further provides compositions comprising one ormore compounds of formula (I) or a pharmaceutically acceptable salt,ester or prodrug thereof, and a pharmaceutically acceptable carrier ordiluent. It will be appreciated that the compounds of formula (I)) inthis invention may be derivatized at functional groups to provideprodrug derivatives which are capable of conversion back to the parentcompounds in vivo. Examples of such prodrugs include the physiologicallyacceptable and metabolically labile ester derivatives, such asmethoxymethyl esters, methylthiomethyl esters, or pivaloyloxymethylesters derived from a hydroxyl group of the compound or a carbamoylmoiety derived from an amino group of the compound. Additionally, anyphysiologically acceptable equivalents of the compounds of formula (I),similar to metabolically labile esters or carbamates, which are capableof producing the parent compounds of formula (I) in vivo, are within thescope of this invention.

As used herein, the term “pharmaceutically acceptable salts” refers toany acid or base addition salt whose counter-ions are non-toxic to thepatient in pharmaceutical doses of the salts. A host of pharmaceuticallyacceptable salts are well known in the pharmaceutical field. Ifpharmaceutically acceptable salts of the compounds of this invention areutilized in these compositions, those salts are preferably derived frominorganic or organic acids and bases. Included among such acid salts arethe following: acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphor sulfonate,cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate,fumarate, lucoheptanoate, glycerophosphate, hemisulfate, heptanoate,hexanoate, hydrochloride, hydrobromide, hydroiodide,2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate,2-naphthalenesulfonate, nicotinate, oxalate, pamoate, pectinate,persulfate, 3-phenyl-propionate, picrate, pivalate, propionate,succinate, tartrate, thiocyanate, tosylate, undecanoate, hydrohalides(e.g., hydrochlorides and hydrobromides), sulphates, phosphates,nitrates, sulphamates, malonates, salicylates,methylene-bis-b-hydroxynaphthoates, gentisates, isethionates,di-p-toluoyltartrates, ethanesulphonates, cyclohexylsulphamates,quinates, and the like. Pharmaceutically acceptable base addition saltsinclude, without limitation, those derived from alkali or alkaline earthmetal bases or conventional organic bases, such as triethylamine,pyridine, piperidine, morpholine, N-methylmorpholine, ammonium salts,alkali metal salts, such as sodium and potassium salts, alkaline earthmetal salts, such as calcium and magnesium salts, salts with organicbases, such as dicyclohexylamine salts, N-methyl-D-glucamine, and saltswith amino acids such as arginine, lysine, and so forth.

Furthermore, the basic nitrogen-containing groups may be quaternizedwith agents like lower alkyl halides, such as methyl, ethyl, propyl andbutyl chlorides, bromides and iodides; dialkyl sulfates, such asdimethyl, diethyl, dibutyl and diamyl sulfates, long chain halides, suchas decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides;aralkyl halides, such as benzyl and phenethyl bromides and others. Wateror oil-soluble or dispersible products are thereby obtained.

The compounds utilized in the compositions and methods of this inventionmay also be modified by appending appropriate functionalities to enhanceselective biological properties. Such modifications are known in the artand include those which increase biological penetration into a givenbiological system (e.g., blood, lymphatic system, central nervoussystem, etc.), increase oral availability, increase solubility to allowadministration by injection, alter metabolism and alter rate ofexcretion.

The pharmaceutical compositions of the invention can be manufactured bymethods well known in the art such as conventional granulating, mixing,dissolving, encapsulating, lyophilizing, or emulsifying processes, amongothers. Compositions may be produced in various forms, includinggranules, precipitates, or particulates, powders, including freezedried, rotary dried or spray dried powders, amorphous powders, tablets,capsules, syrup, suppositories, injections, emulsions, elixirs,suspensions or solutions. Formulations may optionally containstabilizers, pH modifiers, surfactants, bioavailability modifiers andcombinations of these.

The term “unit dosage form” refers to physically discrete units suitableas unitary dosages for human subjects and other mammals, each unitcontaining a predetermined quantity of drug calculated to produce thedesired onset, tolerability, and/or therapeutic effects, in associationwith a suitable pharmaceutical excipient (e.g., an ampoule). Inaddition, more concentrated compositions may be prepared, from which themore dilute unit dosage compositions may then be produced. The moreconcentrated compositions thus will contain substantially more than,e.g., at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more times the amountof one or more syk and/or JAK inhibitors.

Methods for preparing such dosage forms are known to those skilled inthe art (see, for example, REMINGTON'S PHARMACEUTICAL SCIENCES, 18THED., Mack Publishing Co., Easton, Pa. (1990)). In addition,pharmaceutically acceptable salts of the syk and/or JAK inhibitors ofthe present invention (e.g., acid addition salts) may be prepared andincluded in the compositions using standard procedures known to thoseskilled in the art of synthetic organic chemistry and described, e.g.,by J. March, Advanced Organic Chemistry: Reactions, Mechanisms andStructure, 4^(th) Ed. (New York: Wiley-Interscience, 1992).

The compositions typically include a conventional pharmaceutical carrieror excipient and may additionally include other medicinal agents,carriers, adjuvants, diluents, tissue permeation enhancers,solubilizers, and the like. Preferably, the composition will containabout 0.01% to about 90%, preferably about 0.1% to about 75%, morepreferably about 0.1% to 50%, still more preferably about 0.1% to 10% byweight of one or more syk and/or JAK inhibitors, with the remainderconsisting of suitable pharmaceutical carrier and/or excipients.Appropriate excipients can be tailored to the particular composition androute of administration by methods well known in the art, e.g.,REMINGTON'S PHARMACEUTICAL SCIENCES, supra.

Pharmaceutically acceptable carriers that may be used in thesecompositions include ion exchangers, alumina, aluminum stearate,lecithin, serum proteins, such as human serum albumin, buffersubstances, such as phosphates, glycine, sorbic acid, potassium sorbate,partial glyceride mixtures of saturated vegetable fatty acids, water,salts or electrolytes, such as protamine sulfate, disodium hydrogenphosphate, potassium hydrogen phosphate, sodium chloride, zinc salts,colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone,cellulose-based substances, polyethylene glycol, sodiumcarboxymethylcellulose, polyacrylates, waxes,polyethylene-polyoxypropylene-block polymers, polyethylene glycol andwool fat.

Examples of suitable excipients include, but are not limited to,lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia,calcium phosphate, alginates, tragacanth, gelatin, calcium silicate,microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water,saline, syrup, methylcellulose, ethylcellulose,hydroxypropylmethylcellulose, and polyacrylic acids such as Carbopols.The compositions can additionally include lubricating agents such astalc, magnesium stearate, and mineral oil; wetting agents; emulsifyingagents; suspending agents; preserving agents such as methyl-, ethyl-,and propyl-hydroxy-benzoates; pH adjusting agents such as inorganic andorganic acids and bases; sweetening agents; and flavoring agents.

Administration of a composition comprising one or more syk and/or JAKinhibitors with one or more suitable pharmaceutical excipients asadvantageous can be carried out via any of the accepted modes ofadministration. Thus, administration can be, for example, oral, topical,intravenous, subcutaneous, transcutaneous, transdermal, intramuscular,intra joint, parenteral, intra-arteriole, intradermal, intraventricular,intracranial, intraperitoneal, intralesional, intranasal, rectal,vaginal, by inhalation or via an implanted reservoir. The term“parenteral” as used herein includes subcutaneous, intravenous,intramuscular, intra-articular, intra-synovial, intrasternal,intrathecal, intrahepatic, intralesional and intracranial injection orinfusion techniques. Preferably, the compositions are administeredorally or intravenously. The formulations of the invention may bedesigned as short-acting, fast-releasing, or long-acting. Still further,compounds can be administered in a local rather than systemic means,such as administration (e.g., injection) as a sustained releaseformulation. According to a representative embodiment, the compositionsof this invention are formulated for pharmaceutical administration to amammal, preferably a human being.

The compositions of the present invention containing one or more sykand/or JAK inhibitors can be administered repeatedly, e.g., at least 2,3, 4, 5, 6, 7, 8, or more times, or the composition may be administeredby continuous infusion. Suitable sites of administration include, butare not limited to, skin, bronchial, gastrointestinal, anal, vaginal,eye, and ear. The formulations may take the form of solid, semi-solid,lyophilized powder, or liquid dosage forms, such as, for example,tablets, pills, capsules, powders, solutions, suspensions, emulsions,suppositories, retention enemas, creams, ointments, lotions, gels,aerosols, or the like, preferably in unit dosage forms suitable forsimple administration of precise dosages.

The pharmaceutical compositions of this invention may be in any orallyacceptable dosage form, including tablets, capsules, cachets, emulsions,suspensions, solutions, syrups, elixirs, sprays, boluses, lozenges,powders, granules, and sustained-release formulations. Suitableexcipients for oral administration include pharmaceutical grades ofmannitol, lactose, starch, magnesium stearate, sodium saccharine,talcum, cellulose, glucose, gelatin, sucrose, magnesium carbonate, andthe like. In the case of tablets for oral use, carriers that arecommonly used include lactose and corn starch. Lubricating agents, suchas magnesium stearate, are also typically added. For a capsule form,useful diluents include lactose and dried cornstarch. When aqueoussuspensions are required for oral use, the active ingredient is combinedwith emulsifying and suspending agents. If desired, certain sweetening,flavoring or coloring agents may also be added.

In some embodiments, the compositions take the form of a pill, tablet,or capsule, and thus, the composition can contain, along with one ormore syk and/or JAK inhibitors, a diluent such as lactose, sucrose,dicalcium phosphate, and the like; a disintegrant such as starch orderivatives thereof; a lubricant such as magnesium stearate and thelike; and/or a binder such a starch, gum acacia, polyvinylpyrrolidone,gelatin, cellulose and derivatives thereof. A tablet can be made by anycompression or molding process known to those of skill in the art.Compressed tablets may be prepared by compressing in a suitable machinethe syk and/or JAK inhibitors in a free-flowing form, e.g., a powder orgranules, optionally mixed with accessory ingredients, e.g., binders,lubricants, diluents, disintegrants, or dispersing agents. Moldedtablets can be made by molding in a suitable machine a mixture of thepowdered syk and/or JAK inhibitors with any suitable carrier.

Alternatively, the pharmaceutical compositions of this invention may bein the form of suppositories for rectal administration. These may beprepared by mixing the agent with a suitable non-irritating excipientwhich is solid at room temperature but liquid at rectal temperature andtherefore will melt in the rectum to release the drug. Such materialsinclude cocoa butter, beeswax, polyethylene glycol (PEG), hard fat,and/or hydrogenated cocoglyceride. Compositions suitable for rectaladministration may also comprise a rectal enema unit containing one ormore syk and/or JAK inhibitors and pharmaceutically-acceptable vehicles(e.g., 50% aqueous ethanol or an aqueous salt solution) that arephysiologically compatible with the rectum and/or colon. The rectalenema unit contains an applicator tip protected by an inert cover,preferably comprised of polyethylene, lubricated with a lubricant suchas white petrolatum, and preferably protected by a one-way valve toprevent back-flow of the dispensed formula. The rectal enema unit isalso of sufficient length, preferably two inches, to be inserted intothe colon via the anus.

Liquid compositions can be prepared by dissolving or dispersing one ormore syk and/or JAK inhibitors and optionally one or morepharmaceutically acceptable adjuvants in a carrier such as, for example,aqueous saline, aqueous dextrose, glycerol, ethanol, and the like, toform a solution or suspension, e.g., for oral, topical, or intravenousadministration. Pharmaceutical formulations may be prepared as liquidsuspensions or solutions using a sterile liquid, such as oil, water,alcohol, and combinations thereof. Pharmaceutically suitablesurfactants, suspending agents or emulsifying agents, may be added fororal or parenteral administration. Suspensions may include oils, such aspeanut oil, sesame oil, cottonseed oil, corn oil and olive oil.Suspension preparation may also contain esters of fatty acids, such asethyl oleate, isopropyl myristate, fatty acid glycerides and acetylatedfatty acid glycerides. Suspension formulations may include alcohols,such as ethanol, isopropyl alcohol, hexadecyl alcohol, glycerol andpropylene glycol. Ethers, such as poly(ethyleneglycol), petroleumhydrocarbons, such as mineral oil and petrolatum, and water may also beused in suspension formulations.

The pharmaceutical compositions of this invention may also be in atopical form, especially when the target of treatment includes areas ororgans readily accessible by topical application, including diseases ofthe eye, the skin, or the lower intestinal tract. Suitable topicalformulations are readily prepared for each of these areas or organs. Fortopical administration, the composition containing one or more sykand/or JAK inhibitors can be in the form of emulsions, lotions, gels,foams, creams, jellies, solutions, suspensions, ointments, andtransdermal patches.

Topical application for the lower intestinal tract may be effected in arectal suppository formulation (see above) or in a suitable enemaformulation. Topically-transdermal patches may also be used. For topicalapplications, the pharmaceutical compositions may be formulated in asuitable ointment containing the active component suspended or dissolvedin one or more carriers. Carriers for topical administration of thecompounds of this invention include, but are not limited to, mineraloil, liquid petrolatum, white petrolatum, propylene glycol,polyoxyethylene, polyoxypropylene compound, emulsifying wax and water.Alternatively, the pharmaceutical compositions may be formulated in asuitable lotion or cream containing the active components suspended ordissolved in one or more pharmaceutically acceptable carriers. Suitablecarriers include mineral oil, sorbitan monostearate, polysorbate 60,cetyl esters, wax, cetyl alcohol, 2-octyldodecanol, benzyl alcohol andwater.

The pharmaceutical compositions of this invention may also beadministered by nasal aerosol or inhalation. For delivery by inhalation,the compositions can be delivered as a dry powder or in liquid form viaa nebulizer. Such compositions are prepared according to techniquesknown in the art of pharmaceutical formulation and may be prepared assolutions in saline, employing benzyl alcohol or other suitablepreservatives, absorption promoters to enhance bioavailability,fluorocarbons and/or other conventional solubilizing or dispersingagents.

For ophthalmic use, the pharmaceutical compositions may be formulated asmicronized suspensions in isotonic, pH adjusted sterile saline, or,preferably, as solutions in isotonic, pH adjusted sterile saline, eitherwith our without a preservative, such as benzylalkonium chloride.Alternatively, for ophthalmic uses, the pharmaceutical compositions maybe formulated in an ointment, such as petrolatum.

For parenteral administration, the compositions can be in the form ofsterile injectable solutions and sterile packaged powders. Preferably,injectable solutions are formulated at a pH of about 4.5 to about 7.5.

Sterile injectable forms of the compositions of this invention may beaqueous or oleaginous suspension. These suspensions may be formulatedaccording to techniques known in the art using suitable dispersing orwetting agents and suspending agents. The sterile injectable preparationmay also be a sterile injectable solution or suspension in a non-toxicparenterally acceptable diluent or solvent, for example as a solution in1,3-butanediol. Among the acceptable vehicles and solvents that may beemployed are water, Ringer's solution and isotonic sodium chloridesolution. In addition, sterile, fixed oils are conventionally employedas a solvent or suspending medium. For this purpose, any bland fixed oilmay be employed including synthetic mono- or di-glycerides. Fatty acids,such as oleic acid and its glyceride derivatives are useful in thepreparation of injectables, as are natural pharmaceutically-acceptableoils, such as olive oil or castor oil, especially in theirpolyoxyethylated versions. These oil solutions or suspensions may alsocontain a long-chain alcohol diluent or dispersant, such ascarboxymethyl cellulose or similar dispersing agents which are commonlyused in the formulation of pharmaceutically acceptable dosage formsincluding emulsions and suspensions. Other commonly used surfactants,such as Tweens, Spans and other emulsifying agents or bioavailabilityenhancers which are commonly used in the manufacture of pharmaceuticallyacceptable solid, liquid, or other dosage forms may also be used for thepurposes of formulation. Compounds may be formulated for parenteraladministration by injection such as by bolus injection or continuousinfusion. A unit dosage form for injection may be in ampoules or inmulti-dose containers.

The compositions of the present invention can also be provided in alyophilized form. Such compositions may include a buffer, e.g.,bicarbonate, for reconstitution prior to administration, or the buffermay be included in the lyophilized composition for reconstitution with,e.g., water. The lyophilized composition may further comprise a suitablevasoconstrictor, e.g., epinephrine. The lyophilized composition can beprovided in a syringe, optionally packaged in combination with thebuffer for reconstitution, such that the reconstituted composition canbe immediately administered to a patient.

Any of the above dosage forms containing effective amounts are withinthe bounds of routine experimentation and within the scope of theinvention. A therapeutically effective dose may vary depending upon theroute of administration and dosage form. The representative compound orcompounds of the invention is a formulation that exhibits a hightherapeutic index. The therapeutic index is the dose ratio between toxicand therapeutic effects which can be expressed as the ratio between LD₅₀and ED₅₀. The LD₅₀ is the dose lethal to 50% of the population and theED₅₀ is the dose therapeutically effective in 50% of the population. TheLD₅₀ and ED₅₀ are determined by standard pharmaceutical procedures inanimal cell cultures or experimental animals.

Besides those representative dosage forms described above,pharmaceutically acceptable excipients and carriers and dosage forms aregenerally known to those skilled in the art and are included in theinvention. It should be understood that a specific dosage and treatmentregimen for any particular patient will depend upon a variety offactors, including the activity of the specific compound employed, theage, body weight, general health, sex and diet of the patient, and thetime of administration, rate of excretion, drug combination, judgment ofthe treating physician and severity of the particular disease beingtreated. The amount of active ingredient(s) will also depend upon theparticular compound and other therapeutic agent, if present, in thecomposition.

e. Methods of Use

The invention provides methods of inhibiting or decreasing syk and/orJAK activity as well as treating or ameliorating a syk and/or JAKassociated state, symptom, condition, disorder or disease in a patientin need thereof (e.g., human or non-human). In one embodiment, the sykand/or JAK associated state, symptom, condition, disorder or disease ismediated, at least in part by syk and/or JAK kinase activity. In morespecific embodiments, the present invention provides a method fortreating a condition or disorder mediated at least in part by syk and/orJAK kinase activity is cardiovascular disease, inflammatory disease orautoimmune disease.

In one embodiment, the invention provides methods for preventing ortreating a condition in a mammal characterized by undesired thrombosiscomprising the step of administering to the mammal a therapeuticallyeffective amount of a compound of the present invention. Such conditionsinclude, but are not limited, to restenosis, acute coronary syndrome,myocardial infarction, unstable angina, refractory angina, occlusivecoronary thrombosis occurring post-thrombolytic therapy or post-coronaryangioplasty, a thrombotically mediated cerebrovascular syndrome, embolicstroke, thrombotic stroke, transient ischemic attacks, venousthrombosis, deep venous thrombosis, pulmonary embolism, coagulopathy,disseminated intravascular coagulation, thrombotic thrombocytopenicpurpura, thromboangiitis obliterans, thrombotic disease associated withheparin-induced thrombocytopenia, thrombotic complications associatedwith extracorporeal circulation, thrombotic complications associatedwith instrumentation such as cardiac or other intravascularcatheterization, intra-aortic balloon pump, coronary stent or cardiacvalve, conditions requiring the fitting of prosthetic devices, and thelike.

In a further embodiment, the present invention provides a method fortreating thrombosis, immune thrombocytic purura, heparin inducedthrombocytopenia, dilated cardiomypathy, sickle cell disease,atherosclerosis, myocardial infarction, vacular inflammation, unstableangina or acute coronary syndromes.

In another embodiment, the present invention also provides a method fortreating allergy, asthma, theumatoid arthritis, B Cell mediated diseasesuch as Non-Hodgkin's Lymphoma, anti phospholipids syndrome, lupus,psoriasis, multiple sclerosis, end stage renal disease or chroniclymphocytic leukemia.

In another embodiment, the present invention provides a method fortreating hemolytic anemia or immune thrombocytopenic purpura.

The compounds described herein are also potent and/or selectiveinhibitors of JAK kinases. As a consequence of this activity, thecompounds can be used in a variety of in vitro, in vivo, and ex vivocontexts to regulate or inhibit JAK kinase activity, signaling cascadesin which JAK kinases play a role, and the biological responses effectedby such signaling cascades. For example, in one embodiment, thecompounds can be used to inhibit JAK kinase, either in vitro or in vivo,in virtually any cell type expressing the JAK kinase, such as inhematopoietic cells in which, for example, JAK3 is predominantlyexpressed. They may also be used to regulate signal transductioncascades in which JAK kinases, particularly JAK3, play a role. SuchJAK-dependent signal transduction cascades include, but are not limitedto, the signaling cascades of cytokine receptors that involve the commongamma chain, such as, for example, the IL-4, IL-7, IL-5, IL-9, IL-15 andIL-21, or IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 receptor signalingcascades. The compounds may also be used in vitro or in vivo toregulate, and in particular to inhibit, cellular or biological responsesaffected by such JAK-dependent signal transduction cascades. Suchcellular or biological responses include, but are not limited to,IL-4/ramos CD23 upregulation and IL-2 mediated T-cell proliferation.Importantly, the compounds can be used to inhibit JAK kinases in vivo asa therapeutic approach towards the treatment or prevention of diseasesmediated, either wholly or in part, by a JAK kinase activity (referredto herein as “JAK kinase mediated diseases”). Non-limiting examples ofJAK kinase mediated diseases that can be treated or prevented with thecompounds include, but are not limited to, the following: allergies;asthma; autoimmune diseases such as transplant rejection (e.g., kidney,heart, lung, liver, pancreas, skin, small intestine, large intestine,host versus graft reaction (HVGR), and graft versus host reaction(GVHR)), rheumatoid arthritis, and amyotrophic lateral sclerosis; T-cellmediated autoimmune diseases such as multiple sclerosis, psoraiasis, andSjogren's syndrome; Type II inflammatory diseases such as vascularinflammation (including vasculitis, arteritis, atherosclerosis, andcoronary artery disease); diseases of the central nervous system such asstroke; pulmonary diseases such as bronchitis obliteraus and primarypulmonary hypertension; solid, delayed Type IV hypersensitivityreactions; and hematologic malignancies such as leukemia and lymphomas.

Examples of diseases that are mediated, at least in part, by JAK kinasesthat can be treated or prevented according to the methods include, butare not limited to, allergies, asthma, autoimmune diseases such astransplant rejection (e.g., kidney, heart, lung, liver, pancreas, skin,host versus graft reaction (HVGR), etc.), rheumatoid arthritis, andamyotrophic lateral sclerosis, multiple sclerosis, psoraiasis andSjogren's syndrome, Type II inflammatory disease such as vascularinflammation (including vasculitis, ateritis, atherosclerosis andcoronary artery disease) or other inflammatory diseases such asosteoarthritis, inflammatory bowel disease, ulcerative colitis, Crohn'sdisease, idiopathic inflammatory bowel disease, irritable bowelsyndrome, spastic colon, low grade scarring (e.g., scleroderma,increased fibrosis, keloids, post-surgical scars, pulmonary fibrosis,vascular spasms, migraine, reperfusion injury and post myocardialinfarction), and sicca complex or syndrome, diseases of the centralnervous system such as stroke, pulmonary diseases such as bronchitisobliterous and primary and primary pulmonary hypertension, delayed orcell-mediated, Type IV hypersensitivity and solid and hematologicmalignancies such as leukemias and lyphomas.

In another embodiment, this invention provides a method of inhibiting anactivity of a JAK kinase, comprising contacting the JAK kinase with anamount of a compound effective to inhibit an activity of the JAK kinase,wherein the compound is selected from the compounds of this invention.In certain embodiments of the methods described herein, the method iscarried out in vivo.

In another embodiment, this invention provides a method of inhibiting anactivity of a JAK kinase, comprising contacting in vitro a JAK3 kinasewith an amount of a compound effective to inhibit an activity of the JAKkinase, wherein the compound is selected from the compounds of thisinvention.

In a specific embodiment, the compounds can be used to treat and/orprevent rejection in organ and/or tissue transplant recipients (i.e.,treat and/or prevent allorgraft rejection). Allografts can be rejectedthrough either a cell-mediated or humoral immune reaction of therecipient against transplant (histocompability) antigens present on themembranes of the donor's cells. The strongest antigens are governed by acomplex of genetic loci termed human leukocyte group A (HLA) antigens.Together with the ABO blood groups antigens, they are the chieftransplantation antigens detectable in humans.

Rejection following transplantation can generally be broken into threecategories: hyperacute, occurring hours to days followingtransplantation; acute, occurring days to months followingtransplantation; and chronic, occurring months to years followingtransplantation.

Hyperacute rejection is caused mainly by the production of hostantibodies that attack the graft tissue. In a hyperacute rejectionreaction, antibodies are observed in the transplant vascular very soonafter transplantation. Shortly thereafter, vascular clotting occurs,leading to ischemia, eventual necrosis and death. The graft infarctionis unresponsive to known immunosuppressive therapies. Because HLAantigens can be identified in vitro, pre-transplant screening is used tosignificantly reduce hyperacute rejection. As a consequence of thisscreening, hyperacute rejection is relatively uncommon today.

Acute rejection is thought to be mediated by the accumulation of antigenspecific cells in the graft tissue. The T-cell-mediated immune reactionagainst these antigens (i.e., HVGR or GVHR) is the principle mechanismof acute rejection. Accumulation of these cells leads to damage of thegraft tissue. It is believed that both CD4+helper T-cells and CD8+cytotoxic T-cells are involved in the process and that the antigen ispresented by donor and host dendritic cells. The CD4+ helper T-cellshelp recruit other effector cells, such as macrophapges and eosinophils,to the graft. Accessing T-cell activation signal transduction cascades(for example, CD28, CD40L, and CD2 cascades) are also involved.

The cell-mediated acute rejection can be reversed in many cases byintensifying immunotherapy. After successful reversal, severely damagedelements of the graft heal by fibrosis and the remainder of the graftappears normal. After resolution of acute rejection, dosages ofimmunosuppressive drugs can be reduced to very low levels.

Chronic rejection, which is a particular problem in renal transplants,often progresses insidiously despite increased immunosuppressivetherapy. It is thought to be due, in large part, to cell-mediated TypeIV hypersensitivity. The pathologic profile differs from that of acuterejection. The arterial endothelium is primarily involved with extensiveproliferation that may gradually occlude the vessel lumen, leading toischemia, fibrosis, a thickened intima, and atherosclerotic changes.Chronic rejection is mainly due to a progressive obliteration of graftvasculature and resembles a slow, vasculitic process.

In Type IV hypersensitivity, CD8 cytotoxic T-cells and CD4 helper Tcells recognize either intracellular or extracellular synthesizedantigen when it is complexed, respectively, with either Class I or ClassII MHC molecules. Macrophages function as antigen-presenting cells andrelease IL-1, which promotes proliferation of helper T-cells. HelperT-cells release interferon gamma and IL-2, which together regulatedelayed hyperactivity reactions mediated by macrophage activation andimmunity mediated by T cells. In the case of organ transplant, thecytotoxic T-cells destroy the graft cells on contact.

Since JAK kinases play a critical role in the activation of T-cells, thecompounds described herein can be used to treat and/or prevent manyaspects of transplant rejection, and are particularly useful in thetreatment and/or prevention of rejection reactions that are mediated, atleast in part, by T-cells, such as HVGR or GVHR. The compounds can alsobe used to treat and/or prevent chronic rejection in transplantrecipients and, in particular, in renal transplant recipients. Thecompound can also be administered to a tissue or an organ prior totransplanting the tissue or organ in the transplant recipient.

In another embodiment, this invention provides a method of treating aT-cell mediated autoimmune disease, comprising administering to apatient suffering from such an autoimmune disease an amount of acompound effective to treat the autoimmune disease wherein the compoundis selected from the compounds of the invention. In certain embodimentsof the methods the autoimmune disease is multiple sclerosis (MS),psoraisis, or Sjogran's syndrome. Such autoimmune disease include, butare not limited to, those autoimmune diseases that are frequentlydesignated as single organ or single cell-type autoimmune disorders andthose autoimmune disease that are frequently designated as involvingsystemic autoimmune disorder. Non-limiting examples of diseasesfrequently designated as single organ or single cell-type autoimmunedisorders include: Hashimoto's thyroiditis, autoimmune hemolytic anemia,autoimmune atrophic gastritis of pernicious anemia, autoimmuneencephalomyelitis, autoimmune orchitis, Goodpasture's disease,autoimmune thrombocytopenia, sympathetic ophthalmia, myasthenia gravis,Graves' disease, primary biliary cirrhosis, chronic aggressivehepatitis, ulcerative colitis and membranous glomerulopathy.Non-limiting examples of diseases often designated as involving systemicautoimmune disorder include: systemic lupus erythematosis, rheumatoidarthritis, Sjogren's syndrome, Reiter's syndrome,polymyositis-dermatomyositis, systemic sclerosis, polyarteritis nodosa,multiple sclerosis and bullous pemphigoid. Additional autoimmunediseases, which can be .beta.-cell (humoral) based or T-cell based,include Cogan's syndrome, ankylosing spondylitis, Wegener'sgranulomatosis, autoimmune alopecia, Type I or juvenile onset diabetes,and thyroiditis.

The types of autoimmune diseases that may be treated or prevented withsuch prodrugs generally include those disorders involving tissue injurythat occurs as a result of a humoral and/or cell-mediated response toimmunogens or antigens of endogenous and/or exogenous origin. Suchdiseases are frequently referred to as diseases involving thenonanaphylactic (i.e., Type II, Type III and/or Type IV)hypersensitivity reactions.

Type I hypersensitivity reactions generally result from the release ofpharmacologically active substances, such as histamine, from mast and/orbasophil cells following contact with a specific exogenous antigen. Asmentioned above, such Type I reactions play a role in numerous diseases,including allergic asthma, allergic rhinitis, etc.

Type II hypersensitivity reactions (also referred to as cytotoxic,cytolytic complement-dependent or cell-stimulating hypersensitivityreactions) result when immunoglobulins react with antigenic componentsof cells or tissue, or with an antigen or hapten that has becomeintimately coupled to cells or tissue. Diseases that are commonlyassociated with Type II hypersensitivity reactions include, but are notlimited, to autoimmune hemolytic anemia, erythroblastosis fetalis andGoodpasture's disease.

Type III hypersensitivity reactions, (also referred to as toxic complex,soluble complex, or immune complex hypersensitivity reactions) resultfrom the deposition of soluble circulating antigen-immunoglobulincomplexes in vessels or in tissues, with accompanying acute inflammatoryreactions at the site of immune complex deposition. Non-limitingexamples of prototypical Type III reaction diseases include the Arthusreaction, rheumatoid arthritis, serum sickness, systemic lupuserythematosis, certain types of glomerulonephritis, multiple sclerosisand bullous pemphingoid.

Type IV hypersensitivity reactions (frequently called cellular,cell-mediated, delayed, or tuberculin-type hypersensitivity reactions)are caused by sensitized T-lymphocytes which result from contact with aspecific antigen. Non-limiting examples of diseases cited as involvingType IV reactions are contact dermatitis and allograft rejection.

Autoimmune diseases associated with any of the above nonanaphylactichypersensitivity reactions may be treated or prevented with the prodrugsaccording to structural formulae (I) and (Ia). In particular, themethods may be used to treat or prevent those autoimmune diseasesfrequently characterized as single organ or single cell-type autoimmunedisorders including, but not limited to: Hashimoto's thyroiditis,autoimmune hemolytic anemia, autoimmune atrophic gastritis of perniciousanemia, autoimmune encephalomyelitis, autoimmune orchitis, Goodpasture'sdisease, autoimmune thrombocytopenia, sympathetic ophthalmia, myastheniagravis, Graves' disease, primary biliary cirrhosis, chronic aggressivehepatitis, ulcerative colitis and membranous glomerulopathy, as well asthose autoimmune diseases frequently characterized as involving systemicautoimmune disorder, which include but are not limited to: systemiclupus erythematosis (SLE), rheumatoid arthritis, Sjogren's syndrome,Reiter's syndrome, polymyositis-dermatomyositis, systemic sclerosis,polyarteritis nodosa, multiple sclerosis and bullous pemphigoid.

It will be appreciated by skilled artisans that many of the above-listedautoimmune diseases are associated with severe symptoms, theamelioration of which provides significant therapeutic benefit even ininstances where the underlying autoimmune disease may not beameliorated.

Therapy using the compounds described herein can be applied alone, or itcan be applied in combination with or adjunctive to other commonimmunosuppressive therapies, such as, for example, the following:mercaptopurine; corticosteroids such as prednisone; methylprednisoloneand prednisolone; alkylating agents such as cyclophosphamide;calcineurin inhibitors such as cyclosporine, sirolimus, and tacrolimus;inhibitors of inosine monophosphate dehydrogenase (IMPDH) such asmycophenolate, mycophenolate mofetil, and azathioprine; and agentsdesigned to suppress cellular immunity while leaving the recipient'shumoral immunologic response intact, including various antibodies (forexample, antilymphocyte globulin (ALG), antithymocyte globulin (ATG),monoclonal anti-T-cell antibodies (OKT3)) and irradiation. These variousagents can be used in accordance with their standard or common dosages,as specified in the prescribing information accompanying commerciallyavailable forms of the drugs (see also: the prescribing information inthe 2006 Edition of The Physician's Desk Reference), the disclosures ofwhich are incorporated herein by reference. Azathioprine is currentlyavailable from Salix Pharmaceuticals, Inc., under the brand name AZASAN;mercaptopurine is currently available from Gate Pharmaceuticals, Inc.,under the brand name PURINETHOL; prednisone and prednisolone arecurrently available from Roxane Laboratories, Inc.; Methyl prednisoloneis currently available from Pfizer; sirolimus (rapamycin) is currentlyavailable from Wyeth-Ayerst under the brand name RAPAMUNE; tacrolimus iscurrently available from Fujisawa under the brand name PROGRAF;cyclosporine is current available from Novartis under the brand dameSANDIMMUNE and from Abbott under the brand name GENGRAF; IMPDHinhibitors such as mycophenolate mofetil and mycophenolic acid arecurrently available from Roche under the brand name CELLCEPT and fromNovartis under the brand name MYFORTIC; azathioprine is currentlyavailable from Glaxo Smith Kline under the brand name IMURAN; andantibodies are currently available from Ortho Biotech under the brandname ORTHOCLONE, from Novartis under the brand name SIMULECT(basiliximab), and from Roche under the brand name ZENAPAX (daclizumab).

In another embodiment, the compounds could be administered either incombination or adjunctively with an inhibitor of a syk kinase. sykkinase is a tyrosine kinase known to play a critical role in Fcγreceptor signaling, as well as in other signaling cascades, such asthose involving B-cell receptor signaling (Turner et al., (2000),Immunology Today 21:148-154) and integrins beta (1), beta (2), and beta(3) in neutrophils (Mocsai et al., (2002), Immunity 16:547-558). Forexample, syk kinase plays a pivotal role in high affinity IgE receptorsignaling in mast cells that leads to activation and subsequent releaseof multiple chemical mediators that trigger allergic attacks. However,unlike the JAK kinases, which help regulate the pathways involved indelayed or cell-mediated Type IV hypersensitivity reactions, syk kinasehelps regulate the pathways involved in immediate IgE-mediated, Type Ihypersensitivity reactions. Certain compounds that affect the sykpathway may or may not also affect the JAK pathways.

Suitable syk inhibitory compounds are described, for example, in Ser.No. 10/355,543 filed Jan. 31, 2003 (publication no. 2004/0029902); WO03/063794; Ser. No. 10/631,029 filed Jul. 29, 2003; WO 2004/014382; Ser.No. 10/903,263 filed Jul. 30, 2004; PCT/US2004/24716 filed Jul. 30, 2004(WO005/016893); Ser. No. 10/903,870 filed Jul. 30, 2004;PCT/US2004/24920 filed Jul. 30, 2004; Ser. No. 60/630,808 filed Nov. 24,2004; Ser. No. 60/645,424 filed Jan. 19, 2005; and Ser. No. 60/654,620,filed Feb. 18, 2005, the disclosures of which are incorporated herein byreference. The described herein and syk inhibitory compounds could beused alone or in combination with one or more conventional transplantrejection treatments, as described above.

In a specific embodiment, the compounds can be used to treat or preventthese diseases in patients that are either initially non-responsive(resistant) to or that become non-responsive to treatment with a sykinhibitory compound or one of the other current treatments for theparticular disease. The compounds could also be used in combination withsyk inhibitory compounds in patients that are syk-compound resistant ornon-responsive. Suitable syk-inhibitory compounds with which thecompounds can be administered are provided infra.

In another embodiment, this invention provides a method of treating aT-cell mediated autoimmune disease, comprising administering to apatient suffering from such an autoimmune disease an amount of acompound effective to treat the autoimmune disease wherein the compoundis selected from the compounds of the invention, as described herein,and the compound is administered in combination with or adjunctively toa compound that inhibits syk kinase with an IC₅₀ in the range of atleast 10 μM.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient,comprising administering to the transplant recipient an amount of acompound effective to treat or prevent the rejection wherein thecompound is selected from the compounds of the invention, as describedherein. In a further embodiment, the compound is administered to atissue or an organ prior to transplanting the tissue or organ in thetransplant recipient.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the rejection is acute rejection, comprising administering to thetransplant recipient an amount of a compound effective to treat orprevent the rejection, wherein the compound is selected from thecompounds of the invention.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the rejection is chronic rejection, comprising administering tothe transplant recipient an amount of a compound effective to treat orprevent the rejection, wherein the compound is selected from thecompounds of the invention.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the rejection is mediated by HVGR or GVHR, comprisingadministering to the transplant recipient an amount of a compoundeffective to treat or prevent the rejection, wherein the compound isselected from the compounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the allograft transplant is selected from a kidney, a heart, aliver, and a lung, comprising administering to the transplant recipientan amount of a compound effective to treat or prevent the rejection,wherein the compound is selected from the compounds of this invention,as described herein.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the allograft transplant is selected from a kidney, a heart, aliver, and a lung, comprising administering to the transplant recipientan amount of a compound effective to treat or prevent the rejectionwherein the compound is selected from the compounds of the invention, asdescribed herein, in which the compound is administered in combinationwith or adjunctively to another immunosuppressant.

In another embodiment, this invention provides a method of treating orpreventing allograft transplant rejection in a transplant recipient, inwhich the allograft transplant is selected from a kidney, a heart, aliver, and a lung, comprising administering to the transplant recipientan amount of a compound effective to treat or prevent the rejection,wherein the compound is selected from the compounds of the invention, asdescribed herein, in which the compound is administered in combinationwith or adjunctively to another immunosuppressant, in which theimmunosuppressant is selected from cyclosporine, tacrolimus, sirolimus,an inhibitor of IMPDH, mycophenolate, mycophanolate mofetil, ananti-T-Cell antibody, and OKT3.

The compounds described herein are cytokine moderators of IL-4signaling. As a consequence, the compounds could slow the response ofType I hypersensitivity reactions. Thus, in a specific embodiment, thecompounds could be used to treat such reactions and, therefore, thediseases associated with, mediated by, or caused by suchhypersensitivity reactions (for example, allergies), prophylactically.For example, an allergy sufferer could take one or more of the JAKselective compounds described herein prior to expected exposure toallergens to delay the onset or progress of, or eliminate altogether, anallergic response.

When used to treat or prevent such diseases, the compounds can beadministered singly, as mixtures of one or more compounds, or in mixtureor combination with other agents useful for treating such diseasesand/or the symptoms associated with such diseases. The compounds mayalso be administered in mixture or in combination with agents useful totreat other disorders or maladies, such as steroids, membranestabilizers, 5-lipoxygenase (5LO) inhibitors, leukotriene synthesis andreceptor inhibitors, inhibitors of IgE isotype switching or IgEsynthesis, IgG isotype switching or IgG synthesis, beta.-agonists,tryptase inhibitors, aspirin, cyclooxygenase (COX) inhibitors,methotrexate, anti-TNF drugs, anti CD20 antibody, PD4 inhibitors, p38inhibitors, PDE4 inhibitors, and antihistamines, to name a few. Thecompounds can be administered per se in the form of prodrugs or aspharmaceutical compositions, comprising an active compound or prodrug.

In another embodiment, this invention provides a method of treating orpreventing a Type IV hypersensitivity reaction, comprising administeringto a subject an amount of a compound effective to treat or prevent thehypersensitivity reaction, wherein the compound is selected from thecompounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a Type IV hypersensitivity reaction, which is practicalprophylactically, comprising administering to a subject an amount of acompound effective to treat or prevent the hypersensitivity reaction,wherein the compound is selected from the compounds of this invention,as described herein, and is administered prior to exposure to anallergen.

In another embodiment, this invention provides a method of inhibiting asignal transduction cascade in which JAK3 kinase plays a role,comprising contacting a cell expressing a receptor involved in such asignaling cascade with a compound wherein the compound is selected fromthe compounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a JAK kinase-mediated disease, comprising administering to asubject an amount of compound effective to treat or prevent the JAKkinase-mediated disease, wherein the compound is selected from thecompounds of this invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a JAK kinase-mediated disease, in which the JAK-mediateddisease is HVGR or GVHR, comprising administering to a subject an amountof compound effective to treat or prevent the JAK kinase-mediateddisease, wherein the compound is selected from the compounds of theinvention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a JAK kinase-mediated disease, in which the JAK-mediateddisease is acute allograft rejection, comprising administering to asubject an amount of compound effective to treat or prevent the JAKkinase-mediated disease, wherein the compound is selected from thecompounds of the invention, as described herein.

In another embodiment, this invention provides a method of treating orpreventing a syk and/or JAK kinase-mediated disease, in which theJAK-mediated disease is chronic allograft rejection, comprisingadministering to a subject an amount of compound effective to treat orprevent the JAK kinase-mediated disease, wherein the compound isselected from the compounds of the invention, as described herein.

Active compounds of the invention typically inhibit the syk and/orJAK/Stat pathway. The activity of a specified compound as an inhibitorof a syk and/or JAK kinase can be assessed in vitro or in vivo. In someembodiments, the activity of a specified compound can be tested in acellular assay.

“Cell proliferative disorder” refers to a disorder characterized byabnormal proliferation of cells. A proliferative disorder does not implyany limitation with respect to the rate of cell growth, but merelyindicates loss of normal controls that affect growth and cell division.Thus, in some embodiments, cells of a proliferative disorder can havethe same cell division rates as normal cells but do not respond tosignals that limit such growth. Within the ambit of “cell proliferativedisorder” is neoplasm or tumor, which is an abnormal growth of tissue.Cancer refers to any of various malignant neoplasms characterized by theproliferation of cells that have the capability to invade surroundingtissue and/or metastasize to new colonization sites.

Generally, cell proliferative disorders treatable with the compoundsdisclosed herein relate to any disorder characterized by aberrant cellproliferation. These include various tumors and cancers, benign ormalignant, metastatic or non-metastatic. Specific properties of cancers,such as tissue invasiveness or metastasis, can be targeted using themethods described herein. Cell proliferative disorders include a varietyof cancers, including, among others, ovarian cancer, renal cancer,gastrointestinal cancer, kidney cancer, bladder cancer, pancreaticcancer, lung squamous carcinoma, and adenocarcinoma.

In some embodiments, the cell proliferative disorder treated is ahematopoietic neoplasm, which is aberrant growth of cells of thehematopoietic system. Hematopoietic malignancies can have its origins inpluripotent stem cells, multipotent progenitor cells, oligopotentcommitted progenitor cells, precursor cells, and terminallydifferentiated cells involved in hematopoiesis. Some hematologicalmalignancies are believed to arise from hematopoietic stem cells, whichhave the ability for self renewal. For instance, cells capable ofdeveloping specific subtypes of acute myeloid leukemia (AML) (Cynthia K.Hahn, Kenneth N. Ross, Rose M. Kakoza, Steven Karr, Jinyan Du, Shao-EOng, Todd R. Golub, Kimberly Stegmaier, Syk is a new target for AMLdifferentiation, Blood, 2007, 110, Abstract 209) upon transplantationdisplay the cell surface markers of hematopoietic stem cells,implicating hematopoietic stem cells as the source of leukemic cells.Blast cells that do not have a cell marker characteristic ofhematopoietic stem cells appear to be incapable of establishing tumorsupon transplantation (Blaire et al., 1997, Blood 89:3104-3112). The stemcell origin of certain hematological malignancies also finds support inthe observation that specific chromosomal abnormalities associated withparticular types of leukemia can be found in normal cells ofhematopoietic lineage as well as leukemic blast cells. For instance, thereciprocal translocation t(9q34;22q11) associated with approximately 95%of chronic myelogenous leukemia appears to be present in cells of themyeloid, erythroid, and lymphoid lineage, suggesting that thechromosomal aberration originates in hematopoietic stem cells. Asubgroup of cells in certain types of CML displays the cell markerphenotype of hematopoietic stem cells.

Although hematopoietic neoplasms often originate from stem cells,committed progenitor cells or more terminally differentiated cells of adevelopmental lineage can also be the source of some leukemias. Forexample, forced expression of the fusion protein Bcr/Abl (associatedwith chronic myelogenous leukemia) in common myeloid progenitor orgranulocyte/macrophage progenitor cells produces a leukemic-likecondition. Moreover, some chromosomal aberrations associated withsubtypes of leukemia are not found in the cell population with a markerphenotype of hematopoietic stem cells, but are found in a cellpopulation displaying markers of a more differentiated state of thehematopoietic pathway (Turhan et al., 1995, Blood 85:2154-2161). Thus,while committed progenitor cells and other differentiated cells may haveonly a limited potential for cell division, leukemic cells may haveacquired the ability to grow unregulated, in some instances mimickingthe self-renewal characteristics of hematopoietic stem cells (Passegueet al., Proc. Natl. Acad. Sci. USA, 2003, 100:11842-9).

In some embodiments, the hematopoietic neoplasm treated is a lymphoidneoplasm, where the abnormal cells are derived from and/or display thecharacteristic phenotype of cells of the lymphoid lineage. Lymphoidneoplasms can be subdivided into B-cell neoplasms, T and NK-cellneoplasms, and Hodgkin's lymphoma. B-cell neoplasms can be furthersubdivided into precursor B-cell neoplasm and mature/peripheral B-cellneoplasm. Exemplary B-cell neoplasms are precursor B-lymphoblasticleukemia/lymphoma (precursor B-cell acute lymphoblastic leukemia) whileexemplary mature/peripheral B-cell neoplasms are B-cell chroniclymphocytic leukemia/small lymphocytic lymphoma, B-cell prolymphocyticleukemia, lymphoplasmacytic lymphoma, splenic marginal zone B-celllymphoma, hairy cell leukemia, plasma cell myeloma/plasmacytoma,extranodal marginal zone B-cell lymphoma of MALT type, nodal marginalzone B-cell lymphoma, follicular lymphoma, mantle-cell lymphoma, diffuselarge B-cell lymphoma, mediastinal large B-cell lymphoma, primaryeffusion lymphoma, and Burkitt's lymphoma/Burkitt cell leukemia. T-celland Nk-cell neoplasms are further subdivided into precursor T-cellneoplasm and mature (peripheral) T-cell neoplasms. Exemplary precursorT-cell neoplasm is precursor T-lymphoblastic lymphoma/leukemia(precursor T-cell acute lymphoblastic leukemia) while exemplary mature(peripheral) T-cell neoplasms are T-cell prolymphocytic leukemia T-cellgranular lymphocytic leukemia, aggressive NK-cell leukemia, adult T-celllymphoma/leukemia (HTLV-1), extranodal NK/T-cell lymphoma, nasal type,enteropathy-type T-cell lymphoma, hepatosplenic gamma-delta T-celllymphoma, subcutaneous panniculitis-like T-cell lymphoma, Mycosisfungoides/Sezary syndrome, Anaplastic large-cell lymphoma, T/null cell,primary cutaneous type, Peripheral T-cell lymphoma, not otherwisecharacterized, Angioimmunoblastic T-cell lymphoma, Anaplastic large-celllymphoma, T/null cell, primary systemic type. The third member oflymphoid neoplasms is Hodgkin's lymphoma, also referred to as Hodgkin'sdisease. Exemplary diagnosis of this class that can be treated with thecompounds include, among others, nodular lymphocyte-predominantHodgkin's lymphoma, and various classical forms of Hodgkin's disease,exemplary members of which are Nodular sclerosis Hodgkin's lymphoma(grades 1 and 2), Lymphocyte-rich classical Hodgkin's lymphoma, Mixedcellularity Hodgkin's lymphoma, and Lymphocyte depletion Hodgkin'slymphoma. In various embodiments, any of the lymphoid neoplasms that areassociated with aberrant JAK activity can be treated with the syk and/orJAK inhibitory compounds.

In some embodiments, the hematopoietic neoplasm treated is a myeloidneoplasm. This group comprises a large class of cell proliferativedisorders involving or displaying the characteristic phenotype of thecells of the myeloid lineage. Myeloid neoplasms can be subdivided intomyeloproliferative diseases, myelodysplastic/myeloproliferativediseases, myelodysplastic syndromes, and acute myeloid leukemias.Exemplary myeloproliferative diseases are chronic myelogenous leukemia(e.g., Philadelphia chromosome positive (t(9;22)(qq34;q11)), chronicneutrophilic leukemia, chronic eosinophilic leukemia/hypereosinophilicsyndrome, chronic idiopathic myelofibrosis, polycythemia vera, andessential thrombocythemia. Exemplary myelodysplastic/myeloproliferativediseases are chronic myelomonocytic leukemia, atypical chronicmyelogenous leukemia, and juvenile myelomonocytic leukemia. Exemplarymyelodysplastic syndromes are refractory anemia, with ringedsideroblasts and without ringed sideroblasts, refractory cytopenia(myelodysplastic syndrome) with multilineage dysplasia, refractoryanemia (myelodysplastic syndrome) with excess blasts, 5q-syndrome, andmyelodysplastic syndrome. In various embodiments, any of the myeloidneoplasms that are associated with aberrant syk and/or JAK activity canbe treated with the syk and/or JAK inhibitory compounds.

In some embodiments, the compounds can be used to treat Acute myeloidleukemias (AML), which represent a large class of myeloid neoplasmshaving its own subdivision of disorders. These subdivisions include,among others, AMLs with recurrent cytogenetic translocations, AML withmultilineage dysplasia, and other AML not otherwise categorized.Exemplary AMLs with recurrent cytogenetic translocations include, amongothers, AML with t(8;21)(q22;q22), AML1(CBF-alpha)/ETO, Acutepromyelocytic leukemia (AML with t(15;17)(q22;q11-12) and variants,PML/RAR-alpha), AML with abnormal bone marrow eosinophils(inv(16)(p13q22) or t(16;16)(p13;q11), CBFb/MYH11X), and AML with 11q23(MLL) abnormalities. Exemplary AML with multilineage dysplasia are thosethat are associated with or without prior myelodysplastic syndrome.Other acute myeloid leukemias not classified within any definable groupinclude, AML minimally differentiated, AML without maturation, AML withmaturation, Acute myelomonocytic leukemia, Acute monocytic leukemia,Acute erythroid leukemia, Acute megakaryocytic leukemia, Acutebasophilic leukemia, and Acute panmyelosis with myelofibrosis.

“Treating” within the context of the invention means an alleviation ofsymptoms associated with a disorder or disease, or halt of furtherprogression or worsening of those symptoms, or prevention or prophylaxisof the disease or disorder.

The term “mammal” includes organisms which express syk and/or JAK.Examples of mammals include mice, rats, cows, sheep, pigs, goats,horses, bears, monkeys, dogs, cats and, preferably, humans. Transgenicorganisms which express syk and/or JAK are also included in thisdefinition.

The inventive methods comprise administering an effective amount of acompound or composition described herein to a mammal or non-humananimal. As used herein, “effective amount” of a compound or compositionof the invention includes those amounts that antagonize or inhibit sykand/or JAK. An amount which antagonizes or inhibits syk and/or JAK isdetectable, for example, by any assay capable of determining syk and/orJAK activity, including the one described below as an illustrativetesting method. Effective amounts may also include those amounts whichalleviate symptoms of a syk and/or JAK associated disorder treatable byinhibiting syk and/or JAK. Accordingly, “antagonists of syk” or“antagonists of JAK” include compounds which interact with the syk orJAK, respectively, and modulate, e.g., inhibit or decrease, the abilityof a second compound, e.g., another syk or JAK ligand, to interact withthe syk or JAK, respectively. The syk or JAK binding compounds arepreferably antagonists of syk or JAK, respectively. The language “sykbinding compound” and “JAK-binding compound” (e.g., exhibits bindingaffinity to the receptor) includes those compounds which interact withsyk or JAK resulting in modulation of the activity of syk or JAK,respectively. syk and/or JAK binding compounds may be identified usingan in vitro (e.g., cell and non-cell based) or in vivo method. Adescription of in vitro methods are provided below.

The amount of compound present in the methods and compositions describedherein should be sufficient to cause a detectable decrease in theseverity of the disorder, as measured by any of the assays described inthe examples. The amount of syk and/or JAK modulator needed will dependon the effectiveness of the modulator for the given cell type and thelength of time required to treat the disorder. In certain embodiments,the compositions of this invention may further comprise anothertherapeutic agent. When a second agent is used, the second agent may beadministered either as a separate dosage form or as part of a singledosage form with the compounds or compositions of this invention. Whileone or more of the inventive compounds can be used in an application ofmonotherapy to treat a disorder, disease or symptom, they also may beused in combination therapy, in which the use of an inventive compoundor composition (therapeutic agent) is combined with the use of one ormore other therapeutic agents for treating the same and/or other typesof disorders, symptoms and diseases. Combination therapy includesadministration of the two or more therapeutic agents concurrently orsequentially. The agents may be administered in any order.Alternatively, the multiple therapeutic agents can be combined into asingle composition that can be administered to the patient. Forinstance, a single pharmaceutical composition could comprise thecompound or pharmaceutically acceptable salt, ester or prodrug thereofaccording to the formula I, another therapeutic agent (e.g.,methotrexate) or a pharmaceutically acceptable salt, ester or prodrugthereof, and a pharmaceutically acceptable excipient or carrier.

The invention comprises a compound having the formula I, a method formaking an inventive compound, a method for making a pharmaceuticalcomposition from at least one inventive compound and at least onepharmaceutically acceptable carrier or excipient, and a method of usingone or more inventive compounds to treat a variety of disorders,symptoms and diseases (e.g., inflammatory, autoimmune, neurological,neurodegenerative, oncology and cardiovascular), such as RA,osteoarthritis, irritable bowel disease IBD, asthma, chronic obstructivepulmonary disease COPD and MS. The inventive compounds and theirpharmaceutically acceptable salts and/or neutral compositions may beformulated together with a pharmaceutically acceptable excipient orcarrier and the resulting composition may be administered in vivo tomammals, such as men, women and animals, to treat a variety ofdisorders, symptoms and diseases. Furthermore, the inventive compoundscan be used to prepare a medicament that is useful for treating avariety of disorders, symptoms and diseases.

All of the compounds of the present invention are either potentinhibitors of syk and/or JAK kinases, exhibiting IC₅₀s in the respectiveassay in the range of less than 5 μM, with most being in the nanomolar,and several in the sub-nanomolar, range. In some embodiments, thecompounds of the present invention may be “dual” syk/JAK inhibitors inthat they inhibit both syk and JAK kinase to some degree. In otherembodiments, the compounds of the present invention may selectivelyinhibit syk kinase, but not appreciably inhibit one or more JAK kinases.In other embodiments, the compounds of the present invention mayselectively inhibit JAK kinase, but not appreciably inhibit one or moresyk kinases.

f. Kits

Still another aspect of this invention is to provide a kit comprisingseparate containers in a single package, wherein the inventivepharmaceutical compounds, compositions and/or salts thereof are used incombination with pharmaceutically acceptable carriers to treat states,disorders, symptoms and diseases where syk and/or JAK plays a role.

EXAMPLES

The following examples are offered to illustrate, but not to limit, theclaimed invention.

The starting materials and reagents used in preparing these compoundsgenerally are either available from commercial suppliers, such asAldrich Chemical Co., or are prepared by methods known to those skilledin the art following procedures set forth in references such as Fieserand Fieser's Reagents for Organic Synthesis; Wiley & Sons: New York,1967-2004, Volumes 1-22; Rodd's Chemistry of Carbon Compounds, ElsevierScience Publishers, 1989, Volumes 1-5 and Supplementals; and OrganicReactions, Wiley & Sons: New York, 2005, Volumes 1-65.

The starting materials and the intermediates of the synthetic reactionschemes can be isolated and purified if desired using conventionaltechniques, including but not limited to, filtration, distillation,crystallization, chromatography, and the like. Such materials can becharacterized using conventional means, including physical constants andspectral data.

Unless specified to the contrary, the reactions described hereinpreferably are conducted under an inert atmosphere at atmosphericpressure at a reaction temperature range of from about −78° C. to about150° C., more preferably from about 0° C. to about 125° C., and mostpreferably and conveniently at about room (or ambient) temperature,e.g., about 20° C. to about 75° C.

Referring to the examples that follow, compounds of the presentinvention were synthesized using the methods described herein, or othermethods, which are well known in the art.

The compounds and/or intermediates may be characterized by highperformance liquid chromatography (HPLC) using a Waters Alliancechromatography system with a 2695 Separation Module (Milford, Mass.).The analytical columns may be C-18 SpeedROD RP-18E Columns from MerckKGaA (Darmstadt, Germany). Alternately, characterization may beperformed using a Waters Unity (HPLC) system with Waters Acquity HPLCBEH C-18 2.1 mm×15 mm columns. A gradient elution may be used, typicallystarting with 5% acetonitrile/95% water and progressing to 95%acetonitrile over a period of 5 minutes for the Alliance system and 1minute for the Acquity system. All solvents may contain 0.1%trifluoroacetic acid (TFA). Compounds may be detected by ultravioletlight (UV) absorption at either 220 nm or 254 nm. HPLC solvents may befrom EMD Chemicals, Inc. (Gibbstown, N.J.). In some instances, puritymay be assessed by thin layer chromatography (TLC) using glass backedsilica gel plates, such as, for example, EMD Silica Gel 60 2.5 cm×7.5 cmplates. TLC results may be readily detected visually under ultravioletlight, or by employing well known iodine vapor and other variousstaining techniques.

Mass spectrometric analysis may be performed on one of two Agilent 1100series LCMS instruments with acetonitrile/water as the mobile phase. Onesystem may use TFA as the modifier and measure in positive ion mode[reported as MH+, (M+1) or (M+H)+] and the other may use either formicacid or ammonium acetate and measure in both positive [reported as MH⁺,(M+1) or (M+H)⁺] and negative [reported as M−, (M−1) or (M−H)⁻] ionmodes.

Nuclear magnetic resonance (NMR) analysis may be performed on some ofthe compounds with a Varian 400 MHz NMR (Palo Alto, Calif.). Thespectral reference may be either TMS or the known chemical shift of thesolvent.

The purity of some of the invention compounds may be assessed byelemental analysis (Robertson Microlit, Madison, N.J.).

Melting points may be determined on a Laboratory Devices MeI-Tempapparatus (Holliston, Mass.).

Preparative separations may be carried out as needed, using either anSq16x or an Sg100c chromatography system and prepackaged silica gelcolumns all purchased from Teledyne Isco, (Lincoln, Nebr.). Alternately,compounds and intermediates may be purified by flash columnchromatography using silica gel (230-400 mesh) packing material, or byHPLC using a C-18 reversed phase column. Typical solvents employed forthe Isco systems and flash column chromatography may be dichloromethane,methanol, ethyl acetate, hexane, acetone, aqueous hydroxyamine andtriethyl amine. Typical solvents employed for the reverse phase HPLC maybe varying concentrations of acetonitrile and water with 0.1%trifluoroacetic acid.

General Methods

The following synthetic reaction schemes are merely illustrative of somemethods by which the compounds of the present invention can besynthesized, and various modifications to these synthetic reactionschemes can be made and will be suggested to one skilled in the arthaving referred to the disclosure contained in this application.

Example 1N2-(1H-indazol-6-yl)-N4-methyl-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

To a suspension of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (2.0 g, 10.6mmol) in DCM (32 mL) was added NIS (2.4 g, 10.6 mmol) at roomtemperature. After stirring for 1 h, the resulting precipitate wascollected by filtration to give2,4-dichloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (1.8 g).

To a mixture of 2,4-dichloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (1.80 g,5.73 mmol) in DCM (20 ml) was added TsCl (1.09 g, 5.73 mmol) and TEA(1.60 mL, 11.46 mmol), followed by DMAP (70 mg, 0.573 mmol). Afterstirring for 1 h at room temperature, the solution was concentrated, andthe residue was partitioned between EtOAc and H₂O, the organic layer wasseparated, washed with 1N HCl, 5% NaHCO₃, dried and concentrated to give2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (2.0 g).

To a mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(0.6 g, 1.28 mmol) in nBuOH (2 mL) was added methylamine (2M THF, 0.77mL, 1.54 mmol) and DIPEA (0.274 mL, 1.54 mmol). After stirring atambient temperature for 1 h, the resulting precipitate was collected byfiltration to give2-chloro-N-methyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.275 g).

To a mixture of2-chloro-N-methyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.05 g, 0.11 mmol), Pd(PPh₃)₂Cl₂ (0.015 g, 0.022 mmol) andpyridine-4-ylboronic acid (0.03 g, 0.24 mmol) in p-dioxane (0.7 mL) wasadded a solution of Na₂CO₃ (0.035 g, 0.33 mmol) in water (0.3 mL). Afterdegassing, the mixture was heated at 100° C. for 1.5 h. The mixture waspurified by flash column chromatography (Hexane/EtOAc=1:1) to give2-chloro-N-methyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.02 g).

To a mixture of2-chloro-N-methyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.041 g, 0.091 mmol) in nBuOH (0.8 mL) was added 6-aminoindazole (0.024g, 0.18 mmol) and TMSCl (0.03 mL, 0.23 mmol). After heating at 115° C.for 48 h, the mixture was purified by preparative HPLC to giveN²-(1H-indazol-6-yl)-N⁴-methyl-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine(0.011 g), MS (MH 357.1), λ=214.5, 247.5, 306.8.

Example 21-(4-(4-(4-(methylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

To a mixture of2-chloro-N-methyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.044 g, 0.11 mmol) in nBuOH (0.6 mL) was added1-(4-4-aminophenyl)piperazin-1-yl)ethanone (0.047 g, 0.22 mmol) andTMSCl (0.043 mL, 0.33 mmol). After heating at 115° C. for 48 h, themixture was purified by preparative HPLC to give1-(4-(4-(4-(methylamino-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)phenyl)piperizine-1-yl)ethanone(0.015 g), MS (MH 443.3), λ=259.4, 302.1.

Example 3N2-(1H-indazol-6-yl)-N4-methyl-5-(pyridin-3-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

To a mixture of2-chloro-N-methyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.175 g, 0.38 mmol), Pd(PPh₃)₂Cl₂ (0.053 g, 0.076 mmol) andpyridine-3-ylboronic acid (0.102 g, 0.83 mmol) in dioxane (2 mL) wasadded a solution of Na₂CO₃ (0.12 g, 1.14 mmol) in water (1 mL). Afterdegassing, the mixture was heated at 100° C. for 1.5 h. The mixture waspurified by flash column chromatography (Hexane/EtOAc=1:1) to give2-chloro-N-methyl-5-(pyridin-3-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.084 g).

To a mixture of2-chloro-N-methyl-5-(pyridin-3-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.075 g, 0.182 mmol) in nBuOH (1 mL) was added 6-aminoindazole (0.048g, 0.36 mmol) and TMSCl (0.06 mL, 0.46 mmol). After heating at 115° C.for 72 h, the mixture was purified by preparative HPLC to giveN²-(1H-indazol-6-yl)-N⁴-methyl-5-(pyridin-3-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine(0.03 g), MS (MH 357.2), λ=203.9, 246.3, 305.6.

Example 41-(4-(4-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

To a mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(0.414 g, 0.88 mmol) in nBuOH (1.5 mL) was added cyclopropylamine(0.0674 mL, 1.06 mmol) and DIPEA (0.188 mL, 1.06 mmol). After stirringat ambient temperature for 15 h, the resulting precipitate was collectedby filtration to give2-chloro-N-cyclopropyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.3 g).

To a mixture of2-chloro-N-cyclopropyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.3 g, 0.61 mmol), Pd(PPh₃)₂Cl₂ (0.086 g, 0.122 mmol) andpyridine-4-ylboronic acid (0.165 g, 1.342 mmol) in dioxane (3 mL) wasadded a solution of Na₂CO₃ (0.2 g, 1.83 mmol) in water (1.5 mL). Afterdegassing, the mixture was heated at 100° C. for 1.5 h. The mixture waspurified by flash column chromatography (Hexane/EtOAc=1:1) to give2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.241 g).

To a mixture of2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.12 g, 0.27 mmol) in nBuOH (1 mL) was added1-(4-4-aminophenyl)piperazin-1-yl)ethanone (0.12 g, 0.54 mmol) and TMSCl(0.087 mL, 0.68 mmol). After heating at 115° C. for 48 h, the mixturewas purified by preparative HPLC to give1-(4-(4-(4-(cyclopropylamino-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)phenyl)piperizine-1-yl)ethanone(0.015 g), MS (MH 469.4), λ=203.9, 280.7 nm.

Example 54-(4-aminophenyl)-1-(5-carbamoyl-4-(cyclopropylamino)pyrimidin-2-yl)pyridinium

Step 1: To a stirring solution of carboxylic acid 1.1 (85 g, 540 mmol)in thionyl chloride (425 mL) was added pyridine (8.5 mL, 0.11 mmol),slowly. The reaction was stirred at 75° C. overnight at which time itwas concentrated and dried under vacuum to a light yellow powder whichwas used immediately for the next step.

Step 2: The yellow solid from the Step 1 was slowly diluted with 750 mLof ethanol and refluxed overnight. The next day the reaction wasdetermined to be complete by HPLC and then cooled in an ice bath and thesolid filtered and washed with diethyl ether affording the desired ethylester 1.3 as an off-white powder (91 g, 87% for two steps). MS found forC₇H₈N₂O₄ as (M+H)⁺185.0.

Step 3: Ester 1.3 (22 g, 120 mmol) was dissolved in phosphorousoxychloride (60 mL, 600 mmol) and the mixture treated withN,N-diethylaniline (27 mL, 167 mmol) and the mixture heated to 105° C.until the reaction was determined to be complete by HPLC. It was thencooled to rt and slowly added to 1 L of crushed ice resulting in theformation of a beige precipitate which was collected by filtration anddried under vacuum affording the desired dichloride (1.4) as a lightyellow powder (22.5 g, 85%). ¹H NMR (DMSO-d₆, 400 MHz): δ 9.13 (s, 1H),4.37 (q, 2H), 1.32 (t, 3H).

Step 4: Dichloropyrimidine 1.4 (5.9 g, 27 mmol) was dissolved inacetonitrile (50 mL) and treated sequentially with diisopropylamine (5.2mL, 30 mmol) followed by cyclopropyl amine (1.9 g, 27 mmol) and stirredat rt until all starting material had been consumed. The reactionmixture was then diluted with water to a total volume of 150 mL and theprecipitate collected by filtration affording the desired product as alight yellow solid (6.02 g, 87%). ¹H NMR (DMSO-d₆, 400 MHz): δ 8.60 (S,1H), 8.48 (d, 1H), 4.52 (m, 1H), 4.29 (q, 2H), 2.30 (m, 2H), 2.04 (m,2H), 1.73 (m, 2H), 1.30 (t, 3H).

Step 5: Ethyl ester 1.5 (6.02 g, 24 mmol) was diluted with 1,4-dioxane(26 mL) followed by aqueous lithium hydroxide (1.0 M, 26 mL, 26 mmol)and stirred at rt until all starting material had been converted to thecarboxylic acid. The reaction was then diluted with water to a totalvolume of 100 mL and acidified to pH=2 with 6 M HCl. The resultingsuspension was then filtered and dried by aspiration giving 3.51 g ofthe carboxylic acid (64%). ¹H NMR (DMSO-d₆, 400 MHz): δ 8.64 (d, 1H),8.74 (s, 1H), 4.50 (m, 1H), 2.31 (m, 2H), 2.03 (m, 2H), 1.72 (m, 2H).

Step 6: Carboxylic acid 1.6 (3.15 g, 15 mmol) was dissolved inN,N-dimethylformamide (70 mL) and treated with HOBt (3.13 g, 23 mmol)and EDC (4.4 g, 23 mmol). After stirring ca. 25 min ammonia (0.5 M in1,4-dioxane, 72 mL, 36 mmol) was added and the reaction stirredovernight. The following morning the reaction was diluted with water toa total volume of 500 mL and the desired product collected by filtrationaffording 3.62 g (74%) of a light-beige solid. ¹H NMR (DMSO-d₆, 400MHz): δ 9.30 (d, 1H), 8.54 (s, 1H), 8.15 (d, 1H), 8.09 (s, 1H), 7.74 (d,1H), 7.64 (m, 2H), 7.51 (t, 1H), 3.77 (m, 1H), 1.79 (m, 2H), 1.74 (m,2H), 1.53 (m, 1H), 1.41 (m, 1H).

Step 7: Benzotriazolyl ether 1.7 (50 mg, 0.17 mmol), 4-(pyridyl)-aniline(0.26 mmol) and p-toluenesulfonic acid (30 mg, 0.17 mmol) were dilutedwith 1,4-dioxane (5 mL) and stirred at 120° C. until all startingmaterial had been consumed. The reaction was cooled to rt, diluted withwater and directly purified by preparative HPLC affording the desiredproduct, 1, after lyophilization. MS found for C₁₉H₁₉N₆O as (M)⁺347.3.

Example 61-(4-aminophenyl)-3-(5-carbamoyl-4-(cyclopropylamino)pyrimidin-2-yl)-1H-imidazol-3-ium

The above compound was prepared using a procedure similar to thatdescribed in Example 5, Step 7 using 4-(1H-imidazol-1-yl)aniline. MSfound for C₁₇H₁₈N₇O as (M)⁺336.0.

Example 72-(6-amino-7-chloro-1H-indazol-1-yl)-4-(cyclopropylamino)pyrimidine-5-carboxamide

The above compound was prepared using a procedure similar to thatdescribed in Example 5, Step 7 using 7-chloro-1H-indazol-6-amine(synthesized from 6-aminoindazole using N-chlorosuccinimide in onestep). MS found for C₁₅H₁₇N₇OCl as (M+H)⁺344.2, 346.2.

Examples 8 and 91-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamideand1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide

To a mixture of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (1.67 g, 8.88mmol), p-toluenesulfonyl chloride (1.72 g, 9.02 mmol) and triethylamine(2.50 mL, 18.0 mmol) in CH₂C₂ (15 mL), dimethylaminopyridine (30 mg,0.24 mmol) was added. It was stirred at room temperature for 20 h. Waterand CH₂Cl₂ were added. The organic phase was separated, washed with 1NHCl, then with 5% NaHCO₃, dried over Na₂SO₄, concentrated in vacuo togive 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine as a solid (3.03g).

A solution of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (200 mg,0.584 mmol), nipecotamide (75 mg, 0.586 mmol) and triethylamine (0.130mL, 0.934 mmol) in dioxane (5 mL) was stirred at 70° C. for 20 h. Waterand EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo to give1-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide(250 mg).

A mixture of1-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide(125 mg, 0.288 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (100mg, 0.456 mmol) and trimethylsilyl chloride (0.100 mL, 0.79 mmol) inn-BuOH (3 mL) was stirred at 116° C. for 20 h. n-BuOH was removed invacuo. The residue was purified by HPLC to give1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide(12 mg).

To a solution of1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide(12 mg, 0.019 mmol) in MeOH (2 mL), aq. 1N KOH (1 mL) was added. It wasstirred at 60° C. for 2 h. After being concentrated in vacuo, theresidue was acidified with HOAc (1 mL). The mixture was then purified byHPLC to give1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide(2 mg) (MS 421.5 (M+H)) and1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-3-carboxamide(8 mg) (MS 463.6 (M+H)); UV 202.6, 274.6 nm).

Examples 10 and 114-(4-(aminomethyl)piperidin-1-yl)-N-(4-(piperazin-1-yl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amineand1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A solution of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (200 mg,0.584 mmol), 4-N-Boc-aminomethylpiperidine (125 mg, 0.584 mmol) andtriethylamine (0.160 mL, 1.15 mmol) in dioxane (5 mL) was stirred at 70°C. for 20 h. Water and EtOAc were added. The organic phase wasseparated, dried over Na₂SO₄, concentrated in vacuo to give tert-butyl(1-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(303 mg).

A mixture of tert-butyl(1-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(303 mg, 0.583 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (191mg, 0.872 mmol) and trimethylsilyl chloride (0.400 mL, 3.16 mmol) inn-BuOH (6 mL) was stirred at 135° C. for 40 h. n-BuOH was removed invacuo. The residue was purified by HPLC to give1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(60 mg).

To a solution of1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(60 mg, 0.10 mmol) in MeOH (3 mL), aq. 1N KOH (1 mL) was added. It wasstirred at 60° C. for 3 h. After being concentrated in vacuo, theresidue was acidified with HOAc (1 mL). The mixture was then purified byHPLC to give4-(4-(aminomethyl)piperidin-1-yl)-N-(4-(piperazin-1-yl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine(3 mg) (MS 407.5 (M+H); UV 202.6, 269.9 nm) and1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(15 mg) (MS 449.5 (M+H); UV 202.6, 273.4 nm).

Examples 12 and 13(S)-1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)pyrrolidin-3-oland (S)-1-(4-(4-(4-(3hydroxypyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A solution of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (200 mg,0.584 mmol), (S)-3-hydroxypyrrolidine (58 mg, 0.667 mmol) andtriethylamine (0.160 mL, 1.15 mmol) in dioxane (5 mL) was stirred at 70°C. for 20 h. Water and EtOAc were added. The organic phase wasseparated, dried over Na₂SO₄, concentrated in vacuo to give(S)-1-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)pyrrolidin-3-ol(201 mg). MS 393.3 and 395.3 (M+H, Cl pattern).

A mixture of(S)-1-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)pyrrolidin-3-ol(201 mg, 0.512 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (160mg, 0.730 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) inn-BuOH (5 mL) was stirred at 120° C. for 40 h. n-BuOH was removed invacuo. The residue was purified by HPLC to give(S)-1-(4-(4-(4-(3-hydroxypyrrolidin-1-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(80 mg). MS 576.5 (M+H)

To a solution of(S)-1-(4-(4-(4-(3-hydroxypyrrolidin-1-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(80 mg, 0.14 mmol) in MeOH (3 mL), aq. 1N KOH (1 mL) was added. It wasstirred at 60° C. for 4 h. After being concentrated in vacuo, theresidue was acidified with HOAc (1 mL). The mixture was then purified byHPLC to give(S)-1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)pyrrolidin-3-ol(5 mg) (MS 380.5 (M+H)) and(S)-1-(4-(4-(4-(3-hydroxypyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(20 mg) (MS 422.5 (M+H)).

Example 142-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

To a suspension of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (3.00 g,16.0 mmol) in CH₂Cl₂ (30 mL), N-iodosuccinimide (3.60 g, 16.0 mmol) wasadded. It was stirred at room temperature for 20 h. The fine precipitatewas collected, dried on vacuum to give2,4-dichloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (2.80 g). MS 314.1 and316.1 (M+H)

To a suspension of 2,4-dichloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (2.80g, 8.92 mmol) and p-toluenesulfonyl chloride (1.65 g, 8.66 mmol) inCH₂Cl₂ (30 mL), triethylamine (3.0 mL, 21.6 mmol) anddimethylaminopyridine (0.047 g, 0.38 mmol) were added. The mixture wasstirred at room temperature for 2 h. Water and CH₂Cl₂ were added. Theorganic phase was separated, washed with 1N HCl, then with 5% NaHCO₃,dried over Na₂SO₄, concentrated in vacuo to give2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine as a solid (4.00g). MS 468.2 and 470.2 (M+H)

A mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(310 mg, 0.662 mmol), 4-N-Boc-aminomethyl piperidine (142 mg, 0.663mmol) and triethylamine (0.200 mL, 1.44 mmol) in CH₃CN (10 mL) wasstirred at room temperature for 20 h. Water and EtOAc were added. Theorganic phase was separated, washed with 1N HCl, then with 5% NaHCO₃,dried over Na₂SO₄, concentrated in vacuo to give tert-butyl(1-(2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(418 mg). MS 646.5 and 648.4 (M+H)

A mixture of tert-butyl(1-(2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(418 mg, 0.647 mmol), Pd2(dba)₃ (60 mg, 0.065 mmol), dppf (72 mg, 0.130mmol) and Zn(CN)₂ (91 mg, 0.778 mmol) in DMF (5 mL) was stirred at 70°C. for 20 h. DMF was removed in vacuo. The residue was loaded to a flashcolumn, eluted with a gradient of 10-30% EtOAc in hexane to givetert-butyl(1-(2-chloro-5-cyano-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(180 mg).

A mixture of tert-butyl(1-(2-chloro-5-cyano-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(180 mg, 0.330 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (137mg, 0.625 mmol) and trimethylsilyl chloride (0.300 mL, 2.37 mmol) inn-BuOH (5 mL) was stirred at 135° C. for 40 h. n-BuOH was removed invacuo. The residue was purified by HPLC to give the titled compound (25mg). MS 474.5 (M+H); UV 200.8, 277.8 nm.

Example 151-(4-(4-(6-(4-aminomethyl)piperidin-1-yl)-9H-purin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A mixture of 2,6-dichloropurine (189 mg, 1,00 mmol), 4-N-Boc-aminomethylpiperidine (214 mg, 1.00 mmol) and triethylamine (0.300 mL, 2.15 mmol)in CH₃CN (4 mL) was stirred at 70° C. It turned clear solutioninitially, then white precipitates crashed out, which were collected anddried on vacuum to give tert-butyl(1-(2-chloro-9H-purin-6-yl)piperidin-4-yl)methylcarbamate (310 mg). MS367.4 and 369.4 (M+H, Cl pattern)

A mixture of tert-butyl(1-(2-chloro-9H-purin-6-yl)piperidin-4-yl)methylcarbamate (100 mg, 0.272mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (120 mg, 0.548 mmol)and trimethylsilyl chloride (0.300 mL, 2.37 mmol) in n-BuOH (4 mL) wasstirred at 135° C. for 70 h. n-BuOH was removed in vacuo. The residuewas purified by HPLC to give the titled compound (12 mg). MS 450.6(M+H); UV 200.0, 270.8 nm.

Example 16N-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-yl)-1H-indazol-6-amine

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),4-N-Boc-aminomethyl piperidine (214 mg, 1.00 mmol) and triethylamine(0.300 mL, 2.15 mmol) in CH₃CN (4 mL) was stirred at room temperaturefor 20 h. It was concentrated in vacuo. The residue was dissolved innBuOH (6 mL). 2 mL of the nBuOH solution was taken, to which6-aminoindazole (66 mg, 0.49 mmol) was added. The solution was stirredat 116° C. for 20 h. nBuOH was removed in vacuo. The residue wasdissolved in TFA (2 mL). After being stirred at room temperature for 2h, TFA was removed in vacuo. The residue was purified by HPLC to givethe titled compound (40 mg). MS 342.5 (M+H); UV 205.8, 246.8, 276.8 nm.

Example 174-(4-(aminomethyl)piperidin-1-yl)-5-fluoro-N-(3,4,5-trimethoxyphenyl)pyrimidin-2-amine

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),4-N-Boc-aminomethyl piperidine (214 mg, 1.00 mmol) and triethylamine(0.300 mL, 2.15 mmol) in CH₃CN (4 mL) was stirred at room temperaturefor 20 h. It was concentrated in vacuo. The residue was dissolved innBuOH (6 mL). 2 mL of the nBuOH solution was taken, to which3,4,5-trimethoxyaniline (91 mg, 0.50 mmol) was added. The solution wasstirred at 116° C. for 20 h. nBuOH was removed in vacuo. The residue wasdissolved in TFA (2 mL). After being stirred at room temperature for 2h, TFA was removed in vacuo. The residue was purified by HPLC to givethe titled compound (42 mg). MS 392.5 (M+H); UV 223.8, 257.8 nm.

Example 181-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),4-N-Boc-aminomethyl piperidine (214 mg, 1.00 mmol) and triethylamine(0.300 mL, 2.15 mmol) in CH₃CN (4 mL) was stirred at room temperaturefor 20 h. It was concentrated in vacuo. The residue was dissolved innBuOH (6 mL). 2 mL of the nBuOH solution was taken, to which1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (108 mg, 0.49 mmol) wasadded. The solution was stirred at 116° C. for 20 h. nBuOH was removedin vacuo. The residue was dissolved in TFA (2 mL). After being stirredat room temperature for 2 h, TFA was removed in vacuo. The residue waspurified by HPLC to give the titled compound (43 mg). MS 428.6 (M+H); UV206.8, 265.8 nm.

Examples 19 and 20N-(4-(piperazin-1-yl)phenyl)-4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amineand1-(4-(4-(4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A mixture of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (100 mg, 0.531mmol), piperidine (0.060 mL, 0.607 mmol) and triethylamine (0.200 mL,1.43 mmol) in dioxane (4 mL) was stirred at 70° C. for 20 h. Water andEtOAc were added. The organic phase was separated, dried over Na₂SO₄,concentrated in vacuo to give2-chloro-4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine (123 mg). MS237.3 and 239.3 (M+H, Cl pattern)

A mixture of 2-chloro-4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine(123 mg, 0.520 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (200mg, 0.913 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) inn-BuOH (5 mL) was stirred at 120° C. for 20 h. n-BuOH was removed invacuo. The residue was purified by HPLC to giveN-(4-(piperazin-1-yl)phenyl)-4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine(5 mg) (MS 378.5 (M+H)) and1-(4-(4-(4-(piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(31 mg) (MS 420.5 (M+H); UV 205.8, 275.8 nm).

Examples 21 and 22N-(4-(piperazin-1-yl)phenyl)-4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amineand1-(4-(4-(4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A mixture of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (100 mg, 0.531mmol), pyrrolidine (0.050 mL, 0.600 mmol) and triethylamine (0.200 mL,1.43 mmol) in dioxane (4 mL) was stirred at 70° C. for 20 h. Water andEtOAc were added. The organic phase was separated, dried over Na₂SO₄,concentrated in vacuo to give2-chloro-4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine (116 mg). MS223.3 and 225.3 (M+H, Cl pattern)

A mixture of 2-chloro-4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine(116 mg, 0.521 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (200mg, 0.913 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) inn-BuOH (5 mL) was stirred at 120° C. for 20 h. n-BuOH was removed invacuo. The residue was purified by HPLC to giveN-(4-(piperazin-1-yl)phenyl)-4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine(3 mg) (MS 364.5 (M+H)) and1-(4-(4-(4-(pyrrolidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(30 mg) (MS 406.5 (M+H)).

Examples 23 and 241-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamideand1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide

A mixture of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (140 mg, 0.744mmol), isonipecotamide (95 mg, 0.742 mmol) and triethylamine (0.300 mL,2.15 mmol) in dioxane (4 mL) was stirred at 70° C. for 20 h. Theprecipitates were collected and dried on vacuum to give1-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide(205 mg). MS 280.3 and 282.3 (M+H, Cl pattern).

A mixture of1-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide (80mg, 0.28 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (125 mg,0.57 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) in n-BuOH(3 mL) was stirred at 130° C. for 20 h. n-BuOH was removed in vacuo. Theresidue was purified by HPLC to give1-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide(4 mg) (MS 421.5 (M+H)) and1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidine-4-carboxamide(6 mg) (MS 463.5 (M+H); UV 200.0, 272.8 nm).

Example 252-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)pyrimidine-5-carboxamide

To a solution of ethyl 2,4-dichloropyrimidine-5-carboxylate (221 mg,1.00 mmol) and DIEA (0.347 mL, 2.00 mmol) in CH₃CN (4 mL), a suspensionof 4-N-Boc-aminomethyl piperidine (214 mg, 1.00 mmol) in CH₃CN (4 mL)was added. The mixture was stirred at room temperature for 20 h. Waterand EtOAc were added. The organic phase was separated, washed with 1NHCl, then with 5% NaHCO₃, dried over Na₂SO₄, concentrated in vacuo togive ethyl4-(4-((tert-butoxycarbonyl)methyl)piperidin-1-yl)-2-chloropyrimidine-5-carboxylate(368 mg). MS 399.5 and 401.5 (M+H, Cl pattern).

To a solution of ethyl4-(4-((tert-butoxycarbonyl)methyl)piperidin-1-yl)-2-chloropyrimidine-5-carboxylate(368 mg, 0.923 mmol) in THF (5 mL), aq. 1M LiOH (2.20 mL, 2.20 mmol) wasadded. It was heated to reflux for 5 h. THF was removed in vacuo. Afterbeing acidified with HOAc (2 mL), the residue was purified by HPLC togive4-(4-((tert-butoxycarbonyl)methyl)piperidin-1-yl)-2-chloropyrimidine-5-carboxylicacid (150 mg). MS 371.4 and 373.4 (M+H, Cl pattern).

To a solution of4-(4-((tert-butoxycarbonyl)methyl)piperidin-1-yl)-2-chloropyrimidine-5-carboxylicacid (150 mg, 0.40 mmol) and HOBt (93 mg, 0.61 mmol) in DMF (2 mL), EDC(116 mg, 0.61 mmol) was added. After being stirred for 90 min, NH₃ (0.5M in dioxane, 4.00 mL, 2.00 mmol) was added. The mixture was stirred atroom temperature for 20 h. Water and EtOAc were added. The organic phasewas separated, washed with 5% NaHCO₃, dried over Na₂SO₄, concentrated invacuo to give tert-butyl(1-(2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-5-carbamoylpyrimidin-4-yl)piperidin-4-yl)methylcarbamate(138 mg). MS 469.6 (M+H).

A mixture of tert-butyl(1-(2-(1H-benzo[d][1,2,3]triazol-1-yloxy)-5-carbamoylpyrimidin-4-yl)piperidin-4-yl)methylcarbamate(138 mg, 0.294 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (84mg, 0.38 mmol) and p-toluenesulfonic acid (56 mg, 0.294 mmol) in dioxane(4 mL) was stirred at 100° C. for 3 h. dioxane was removed in vacuo. Theresidue was dissolved in CH₂C₁₂ (5 mL) and TFA (5 mL). The solution wasstirred at room temperature for 30 min. It was then concentrated invacuo. The residue was purified by HPLC to give the titled compound (15mg). MS 453.6 (M+H); UV 222.8, 263.8 nm.

Example 261-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidine-3-carboxamide

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),nipecotamide (128 mg, 1.00 mmol) and DIEA (0.350 mL, 2.01 mmol) in CH₃CN(4 mL) was stirred at room temperature for 20 h. It was concentrated invacuo. The residue was dissolved in nBuOH (4 mL). 2 mL of the nBuOHsolution was taken, to which 6-aminoindazole (100 mg, 0.75 mmol) wasadded. The solution was stirred at 116° C. for 6 h. nBuOH was removed invacuo. The residue was purified by HPLC to give the titled compound (63mg). MS 356.3 (M+H); UV 206.8, 249.8, 276.8, 291.8 nm.

Example 271-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidine-3-carboxamide

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),nipecotamide (128 mg, 1.00 mmol) and DIEA (0.350 mL, 2.01 mmol) in CH₃CN(4 mL) was stirred at room temperature for 20 h. It was concentrated invacuo. The residue was dissolved in nBuOH (4 mL). 2 mL of the nBuOHsolution was taken, to which 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone(150 mg, 0.68 mmol) was added. The solution was stirred at 116° C. for 6h. nBuOH was removed in vacuo. The residue was purified by HPLC to givethe titled compound (75 mg). MS 442.4 (M+H); UV 208.8, 261.8 nm.

Examples 28 and 29 tert-butyl(1-(5-fluoro-2-(4-(N-methylacetamido)phenylamino)pyrimidin-4-yl)piperidin-4-yl)methylcarbamateandN-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)-N-methylacetamide

A mixture of 2,4-dichloro-5-fluoropyrimidine (334 mg, 2.00 mmol),4-N-Boc-aminomethyl piperidine (428 mg, 2.00 mmol) and DIEA (0.700 mL,4.02 mmol) in CH₃CN (8 mL) was stirred at room temperature for 20 h. Itwas concentrated in vacuo. The residue was dissolved in nBuOH (12 mL). 3mL of the nBuOH solution was taken, to whichN-(4-aminophenyl)-N-methylacetamide (98 mg, 0.60 mmol) was added. Thesolution was stirred at 116° C. for 20 h. nBuOH was removed in vacuo.The residue was purified by HPLC to give tert-butyl(1-(5-fluoro-2-(4-(N-methylacetamido)phenylamino)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(65 mg). MS 473.4 (M+H); UV 205.8, 262.8 nm.

A solution of tert-butyl(1-(5-fluoro-2-(4-(N-methylacetamido)phenylamino)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(60 mg, 0.13 mmol) in TFA (2 mL) was stirred at room temperature for 60min. TFA was removed in vacuo. The residue was purified by HPLC to giveN-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)-N-methylacetamide(25 mg). MS 373.3 (M+H); UV 201.8, 266.8 nm.

Examples 30 and 31 tert-butyl(1-(2-(4-carbamoylphenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)methylcarbamateand4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzamide

A mixture of 2,4-dichloro-5-fluoropyrimidine (334 mg, 2.00 mmol),4-N-Boc-aminomethyl piperidine (428 mg, 2.00 mmol) and DIEA (0.700 mL,4.02 mmol) in CH₃CN (8 mL) was stirred at room temperature for 20 h. Itwas concentrated in vacuo. The residue was dissolved in nBuOH (12 mL). 3mL of the nBuOH solution was taken, to which 4-aminobenzamide (82 mg,0.60 mmol) was added. The solution was stirred at 116° C. for 20 h.nBuOH was removed in vacuo. The residue was purified by HPLC to givetert-butyl(1-(2-(4-carbamoylphenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)methylcarbamate(15 mg). MS 445.4 (M+H); UV 212.0, 282.8 nm.

A solution of tert-butyl(1-(2-(4-carbamoylphenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)methylcarbamate(10 mg, 0.023 mmol) in TFA (2 mL) was stirred at room temperature for 60min. TFA was removed in vacuo. The residue was purified by HPLC to give4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzamide(5 mg) MS 345.3 (M+H); UV 210.0, 281.8 nm.

Example 324-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzenesulfonamide

A mixture of 2,4-dichloro-5-fluoropyrimidine (334 mg, 2.00 mmol),4-N-Boc-aminomethyl piperidine (428 mg, 2.00 mmol) and DIEA (0.700 mL,4.02 mmol) in CH₃CN (8 mL) was stirred at room temperature for 20 h. Itwas concentrated in vacuo. The residue was dissolved in nBuOH (12 mL). 3mL of the nBuOH solution was taken, to which sulfanilamide (103 mg, 0.60mmol) was added. The solution was stirred at 116° C. for 20 h. nBuOH wasremoved in vacuo. The residue was purified by HPLC to give4-(4-(4-(N-tBoc-aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzenesulfonamide(10 mg). MS 481.3 (M+H).

A solution of4-(4-(4-(N-tBoc-aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzenesulfonamide(10 mg, 0.021 mmol) in TFA (2 mL) was stirred at room temperature for 60min. TFA was removed in vacuo. The residue was purified by HPLC to givethe titled compound (5 mg) MS 381.3 (M+H)

Example 336-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

A mixture of 2,4-dichloro-5-fluoropyrimidine (334 mg, 2.00 mmol),4-N-Boc-aminomethyl piperidine (428 mg, 2.00 mmol) and DIEA (0.700 mL,4.02 mmol) in CH₃CN (8 mL) was stirred at room temperature for 20 h. Itwas concentrated in vacuo. The residue was dissolved in nBuOH (12 mL). 3mL of the nBuOH solution was taken, to which6-amino-3,4-dihydroquinolin-2(1H)-one (98 mg, 0.60 mmol) was added. Thesolution was stirred at 116° C. for 20 h. nBuOH was removed in vacuo.The residue was purified by HPLC to give tert-butyl(1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(55 mg). MS 471.4 (M+H).

A solution of tert-butyl(1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(55 mg, 0.12 mmol) in TFA (3 mL) was stirred at room temperature for 60min. TFA was removed in vacuo. The residue was purified by HPLC to givethe titled compound (30 mg) MS 371.3 (M+H); UV 206.8, 274.8 nm.

Example 341-(4-(4-(4-(2-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),2-N-Boc-aminomethyl piperidine (214 mg, 1.00 mmol) and DIEA (0.400 mL,2.30 mmol) in CH₃CN (4 mL) was stirred at room temperature for 20 h. Itwas concentrated in vacuo. The residue was dissolved in nBuOH (6 mL). 3mL of the nBuOH solution was taken, to which1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (142 mg, 0.65 mmol) wasadded. The solution was stirred at 116° C. for 20 h. nBuOH was removedin vacuo. The residue was purified by HPLC to give tert-butyl(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-2-yl)methylcarbamate(56 mg). MS 528.4 (M+H).

A solution of tert-butyl(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-2-yl)methylcarbamate(56 mg, 0.11 mmol) in TFA (3 mL) was stirred at room temperature for 60min. TFA was removed in vacuo. The residue was purified by HPLC to givethe titled compound (35 mg) MS 428.4 (M+H); UV 213.8, 258.8 nm.

Examples 35 and 36 tert-butyl(1-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidin-2-yl)methylcarbamateandN-(4-(2-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-yl)-1H-indazol-6-amine

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),2-N-Boc-aminomethyl piperidine (214 mg, 1.00 mmol) and DIEA (0.400 mL,2.30 mmol) in CH₃CN (4 mL) was stirred at room temperature for 20 h. Itwas concentrated in vacuo. The residue was dissolved in nBuOH (6 mL). 3mL of the nBuOH solution was taken, to which 6-aminoindazole (86 mg,0.65 mmol) was added. The solution was stirred at 116° C. for 20 h.nBuOH was removed in vacuo. The residue was purified by HPLC to givetert-butyl(1-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidin-2-yl)methylcarbamate(41 mg). MS 442.3 (M+H); UV 201.8, 246.8, 281.8 nm.

A solution of tert-butyl(1-(2-(1H-indazol-6-ylamino)-5-fluoropyrimidin-4-yl)piperidin-2-yl)methylcarbamate(41 mg, 0.093 mmol) in TFA (4 mL) was stirred at room temperature for 60min. TFA was removed in vacuo. The residue was purified by HPLC to giveN-(4-(2-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-yl)-1H-indazol-6-amine(25 mg) MS 342.3 (M+H);); UV 201.8, 246.8, 278.8 nm.

Example 371-((1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)methyl)urea

To a suspension of6-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one(26 mg, 0.070 mmol) in CH₃CN (1 mL), an aqueous solution of potassiumcyanate (34 mg, 0.42 mmol) in H₂O (1 mL) was added. The mixture wasstirred at 70° C. for 1 h. Solvents were removed in vacuo. The residuewas purified by HPLC to give the titled compound (8 mg). MS 414.3(M+H);); UV 205.8, 276.8 nm

Example 381-(4-(4-(5-fluoro-4-(4-(hydroxymethyl)piperidin-1-yl)pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),4-piperidinemethanol (115 mg, 1.00 mmol) and DIEA (0.400 mL, 2.30 mmol)in CH₃CN (4 mL) was stirred at room temperature for 2 h. It wasconcentrated in vacuo. The residue was dissolved in nBuOH (6 mL). 3 mLof the nBuOH solution was taken, to which1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (120 mg, 0.55 mmol) wasadded. The solution was stirred at 116° C. for 20 h. nBuOH was removedin vacuo. The residue was purified by HPLC to give the titled compound(53 mg). MS 429.3 (M+H); UV 207.8, 266.8 nm.

Example 396-(5-fluoro-4-(4-(hydroxymethyl)piperidin-1-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

A mixture of 2,4-dichloro-5-fluoropyrimidine (167 mg, 1.00 mmol),4-piperidinemethanol (115 mg, 1.00 mmol) and DIEA (0.400 mL, 2.30 mmol)in CH₃CN (4 mL) was stirred at room temperature for 2 h. It wasconcentrated in vacuo. The residue was dissolved in nBuOH (6 mL). 3 mLof the nBuOH solution was taken, to which6-amino-3,4-dihydroquinolin-2(1H)-one (88 mg, 0.54 mmol) was added. Thesolution was stirred at 116° C. for 20 h. nBuOH was removed in vacuo.The residue was purified by HPLC to give the titled compound (52 mg). MS372.3 (M+H); UV 208.8, 275.8 nm.

The following compound was prepared using a procedure similar to thatdescribed in Example 5 with reagent A in place of cyclobutylamine inStep 4.

Example 402-(1H-indazol-6-ylamino)-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

A solution of2-chloro-N-cyclopropyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(200 mg, 0.409 mmol), Pd2 dba3 (23 mg, 0.025 mmol) and dppf (45 mg,0.081 mmol) in DMF (2 mL) was degassed with Ar before being charged withZn(CN)₂ (28 mg, 0.24 mmol). The mixture was stirred at 70° C. for 2 h.Water and EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo. The residue was purified by a flashsilica gel column, eluted with EtOAc/hexane (0-15%) to give2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(110 mg).

A mixture of2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(109 mg, 0.28 mmol), 6-aminoindazole (75 mg, 0.56 mmol) and TMSCl (0.089mL, 0.70 mmol) in nBuOH (4 mL) was stirred at 116° C. for 18 h. It wasthen concentrated in vacuo. The residue was purified by HPLC to give thetitled compound (6 mg). MS 331.1 (M+H); UV 246.2, 307.9 nm.

Example 415-(1-butoxyethyl)-N4-cyclopropyl-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

A solution of2-chloro-N-cyclopropyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(190 mg, 0.389 mmol) in dioxane (3 mL) was degassed with Ar before beingcharged with tetravinyltin (0.105 mL, 0.576 mmol) and Pd(Ph3P)₄ (41 mg,0.035 mmol). The mixture was stirred at 100° C. for 3 h. Water and EtOAcwere added. The organic phase was separated, dried over Na₂SO₄,concentrated in vacuo. The residue was purified by HPLC to give2-chloro-N-cyclopropyl-7-tosyl-5-vinyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(39 mg).

A mixture of2-chloro-N-cyclopropyl-7-tosyl-5-vinyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(39 mg, 0.10 mmol), 6-aminoindazole (40 mg, 0.30 mmol) and TMSCl (0.050mL, 0.40 mmol) in nBuOH (1 mL) was stirred at 116° C. for 18 h. It wasthen concentrated in vacuo. The residue was purified by HPLC to give5-(1-butoxyethyl)-N4-cyclopropyl-N-2-(1H-indazol-6-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine(20 mg).

To a solution of5-(1-butoxyethyl)-N4-cyclopropyl-N2-(1H-indazol-6-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine(20 mg, 0.036 mmol) in dioxane (2 mL), aq. 1N KOH (1.0 mL, 1.0 mmol) wasadded. The mixture was stirred at 65° C. for 18 h. After beingconcentrated in vacuo, the residue was acidified with HOAc (1 mL); itwas then purified by HPLC to give the titled compound (3 mg). MS 406.2(M+H); UV 203.8, 223.8, 243.8, 315.0 nm.

Example 421-(2-(1H-indazol-6-ylamino)-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-5-yl)ethanone

A solution of2-chloro-N-cyclopropyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(489 mg, 1.00 mmol) in dioxane (6 mL) was degassed with Ar before beingcharged with tributyl(ethoxyvinyl)tin (0.371 mL, 1.10 mmol) andPd(Ph3P)₄ (90 mg, 0.078 mmol). The mixture was stirred at 100° C. for 18h. Water and EtOAc were added. The organic phase was separated, driedover Na₂SO₄, concentrated in vacuo. The residue was purified by HPLC togive1-(2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-5-yl)ethanone(185 mg).

A mixture of1-(2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-5-yl)ethanone(85 mg, 0.21 mmol), 6-aminoindazole (56 mg, 0.42 mmol) and TMSCl (0.070mL, 0.55 mmol) in nBuOH (3 mL) was stirred at 116° C. for 18 h. It wasthen concentrated in vacuo. The residue was purified by HPLC to give thetitled compound (30 mg). MS 348.1 (M+H); UV 248.7, 305.7 nm.

Example 43N4-cyclopropyl-N2-(1H-indazol-6-yl)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

A solution of2-chloro-N-cyclopropyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(225 mg, 0.460 mmol) in dioxane (3 mL) was degassed with Ar before beingcharged with 4-tributylstannylpyridine (371 mg, 1.01 mmol) and Pd(Ph3P)₄(98 mg, 0.085 mmol). The mixture was stirred at 100° C. for 18 h. Waterand EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo. The residue was purified by HPLC to give2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(51 mg).

A mixture of2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(51 mg, 0.12 mmol), 6-aminoindazole (31 mg, 0.23 mmol) and TMSCl (0.040mL, 0.32 mmol) in nBuOH (2 mL) was stirred at 116° C. for 18 h. It wasthen concentrated in vacuo. The residue was purified by HPLC to give thetitled compound (12 mg). MS 383.2 (M+H); UV 204.9, 250.9, 307.9 nm.

Example 441-(4-(4-(4-(piperidin-4-ylmethylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(244 mg, 0.520 mmol), 1-N-BOC-4-aminomethyl piperidine hydrochloride(130 mg, 0.520 mmol) and TEA (0.200 mL, 1.44 mmol) in CH₃CN (6 mL) wasstirred at room temperature for 16 h. Water and EtOAc were added. Theorganic phase was separated, washed with 1N HCl, then with 5% NaHCO₃,dried over Na₂SO₄, concentrated in vacuo to give tert-butyl4-((2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)methyl)piperidine-1-carboxylate(336 mg).

A solution of tert-butyl4-((2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)methyl)piperidine-1-carboxylate(336 mg, 0.520 mmol) in dioxane (4 mL) was degassed with Ar before beingcharged with 4-tributylstannylpyridine (452 mg, 1.23 mmol) and Pd(Ph₃P)₄(110 mg, 0.095 mmol). The mixture was stirred at 100° C. for 4 h. Waterand EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo. The residue was purified by a flashcolumn, eluted with EtOAc/hexane (0-100%) to give tert-butyl4-((2-chloro-5-(pyridine-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)methyl)piperidine-1-carboxylate(90 mg).

A mixture of tert-butyl4-((2-chloro-5-(pyridine-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)methyl)piperidine-1-carboxylate(90 mg, 0.15 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (66 mg,0.30 mmol) and TMSCl (0.100 mL, 0.79 mmol) in nBuOH (4 mL) was stirredat 135° C. for 68 h. It was then concentrated in vacuo. The residue waspurified by HPLC to give the titled compound (26 mg). MS 526.47 (M+H);UV 202.6, 259.2, 304.3 nm.

Example 452-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(piperidin-4-ylmethylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

A solution of tert-butyl4-((2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)methyl)piperidine-1-carboxylate(360 mg, 0.557 mmol), Pd2 dba3 (50 mg, 0.055 mmol) and dppf (62 mg, 0.11mmol) in DMF (5 mL) was degassed with Ar before being charged withZn(CN)₂ (80 mg, 0.68 mmol). The mixture was stirred at 70° C. for 18 h.Water and EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo. The residue was purified by a flashsilica gel column, eluted with EtOAc/hexane (0-35%) to give tert-butyl4-((2-chloro-5-cyano-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)methyl)piperidine-1-carboxylate(127 mg).

A mixture of tert-butyl4-((2-chloro-5-cyano-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)methyl)piperidine-1-carboxylate(127 mg, 0.233 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (100mg, 0.456 mmol) and TMSCl (0.200 mL, 1.58 mmol) in nBuOH (5 mL) wasstirred at 135° C. for 68 h. It was then concentrated in vacuo. Theresidue was purified by HPLC to give the titled compound (25 mg). MS 475(M+H); UV 204.9, 265.1, 297.2 nm.

Example 462-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

A mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(310 mg, 0.662 mmol), 4-N-BOC-aminomethyl piperidine (142 mg, 0.663mmol) and TEA (0.200 mL, 1.44 mmol) in CH₃CN (10 mL) was stirred at roomtemperature for 16 h. Water and EtOAc were added. The organic phase wasseparated, washed with 1N HCl, then with 5% NaHCO₃, dried over Na₂SO₄,concentrated in vacuo to give tert-butyl(1-(2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(418 mg).

A solution of tert-butyl(1-(2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(418 mg, 0.647 mmol), Pd₂ dba₃ (60 mg, 0.065 mmol) and dppf (72 mg, 0.13mmol) in DMF (5 mL) was degassed with Ar before being charged withZn(CN)₂ (91 mg, 0.78 mmol). The mixture was stirred at 70° C. for 18 h.Water and EtOAc were added. The organic phase was separated, dried overNa₂SO₄, concentrated in vacuo. The residue was purified by a flashsilica gel column, eluted with EtOAc/hexane (10-30%) to give tert-butyl(1-(2-chloro-5-cyano-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(180 mg).

A mixture of tert-butyl(1-(2-chloro-5-cyano-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(180 mg, 0.330 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (137mg, 0.625 mmol) and TMSCl (0.300 mL, 2.37 mmol) in nBuOH (5 mL) wasstirred at 135° C. for 48 h. It was then concentrated in vacuo. Theresidue was purified by HPLC to give the titled compound (25 mg). MS474.5 (M+H).

Example 47N-(4-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A mixture of2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(85 mg, 0.19 mmol), N-(4-aminophenyl)-N-methylacetamide (72 mg, 0.44mmol) and TMSCl (0.100 mL, 0.79 mmol) in nBuOH (3 mL) was stirred at135° C. for 18 h. It was then concentrated in vacuo. The residue waspurified by HPLC to give the titled compound (25 mg). MS 414.3 (M+H); UV202.8, 280.8 nm.

Example 486-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

A mixture of2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(85 mg, 0.19 mmol), 6-amino-3,4-dihydroquinolin-2(1H)-one (70 mg, 0.43mmol) and TMSCl (0.100 mL, 0.79 mmol) in nBuOH (3 mL) was stirred at135° C. for 18 h. More TMSCl (0.200 mL) was added. Stirring wascontinued at 135° C. for another 68 h. It was then concentrated invacuo. The residue was purified by HPLC to give the titled compound (5mg). MS 412.3 (M+H); UV 201.8, 295.8 nm.

Example 495-bromo-N4-cyclobutyl-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

To a suspension of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (105 mg,0.560 mmol) in CH₂Cl₂ (7 mL) at room temperature, NBS (109 mg, 0.610mmol) was added. The mixture was stirred at room temperature for 18 h.CH₂Cl₂ was removed in vacuo. The residue was partitioned between waterand EtOAc. The organic phase was separated, washed with 5% NaHCO₃, driedover Na₂SO₄, concentrated in vacuo to give5-bromo-2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (142 mg).

A solution of 5-bromo-2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (142 mg,0.530 mmol), cyclobutylamine (0.090 mL, 1.06 mmol) and TEA (0.150 mL,1.08 mmol) in nBuOH (6 mL) was stirred at 70° C. for 18 h. CH₂Cl₂ andH₂O were added. The organic phase was separated, washed with 5% NaHCO₃,then with 1N HCl, dried over Na₂SO₄, concentrated in vacuo to give5-bromo-2-chloro-N-cyclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (154mg).

A mixture of5-bromo-2-chloro-N-cyclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (64 mg,0.21 mmol), 6-aminoindazole (56 mg, 0.42 mmol) and TMSCl (0.050 mL, 0.40mmol) in nBuOH (2 mL) was stirred at 116° C. for 4 h. It was thenconcentrated in vacuo. The residue was purified by HPLC to give asample, which was further purified by preparative TLC using CH₂Cl₂/MeOH(95/5) as developing solvents to give the titled compound (1 mg). MS398.3, 400.3 (M+H, Br pattern); UV 203.8, 247.4, 309.1 nm.

Example 505-chloro-N4-cyclobutyl-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

To a suspension of2-chloro-N-cyclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (102 mg, 0.460mmol) in CH₂Cl₂ (5 mL) at room temperature, NCS (68 mg, 0.51 mmol) wasadded. The mixture was stirred at reflux for 18 h. After beingconcentrated in vacuo, the residue was purified by HPLC to give2,5-dichloro-N-cyclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (23 mg).

A mixture of2,5-dichloro-N-cyclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (23 mg,0.089 mmol), 6-aminoindazole (40 mg, 0.30 mmol) and TMSCl (0.040 mL,0.32 mmol) in nBuOH (2 mL) was stirred at 116° C. for 18 h. It was thenconcentrated in vacuo. The residue was purified by HPLC to give thetitled compound (8 mg). MS 354.0, 356.0 (M+H, Cl pattern); UV 201.4,246.2, 307.9 nm.

Example 512-(1H-indazol-6-ylamino)-4-(cyclobutylamino)-6,7-dihydropyrrolo[2,3-d]pyrimidin-5-one

A mixture of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (200 mg, 1.06mmol) and SelectFluor (560 mg, 1.58 mmol) in CH₃CN (5 mL) and HOAc (1mL) was stirred at 70° C. for 18 h. The mixture was then concentrated invacuo. The residue was purified by HPLC to give2,4-dichloro-5-fluoro-7H-pyrrolo[2,3-d]pyrimidine (50 mg).

A solution of 2,4-dichloro-5-fluoro-7H-pyrrolo[2,3-d]pyrimidine (50 mg,0.24 mmol), cyclobutylamine (0.041 mL, 0.48 mmol) and TEA (0.070 mL,0.50 mmol) in nBuOH (2 mL) was stirred at 70° C. for 5 h. CH₂Cl₂ and H₂Owere added. The organic phase was separated, washed with 5% NaHCO₃, thenwith 1N HCl, dried over Na₂SO₄, concentrated in vacuo to give2-chloro-N-cyclobutyl-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-4-amine (54mg).

A mixture of2-chloro-N-cyclobutyl-5-fluoro-7H-pyrrolo[2,3-d]pyrimidin-4-amine (54mg, 0.22 mmol), 6-aminoindazole (90 mg, 0.68 mmol) and TMSCl (0.090 mL,0.71 mmol) in nBuOH (3 mL) was stirred at 116° C. for 18 h. It was thenconcentrated in vacuo. The residue was purified by HPLC to give asample, which was further purified by preparative TLC using CH₂Cl₂/MeOH(95/5) as developing solvents to give the titled compound (2 mg). MS336.1 (M+H)

Example 521-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol

To a solution of 4-nitrobenzenesulfonyl chloride (512 mg, 2.31 mmol) inCH₂Cl₂ (8 mL) at room temperature, 4-hydroxypiperidine (404 mg, 4.00mmol) was added. Triethylamine (0.400 mL, 2.88 mmol) was also added. Themixture was stirred at room temperature overnight. Water and EtOAc wereadded. The organic phase was separated, washed with 1N HCl, dried overNa₂SO₄, concentrated in vacuo to give1-(4-nitrophenylsulfonyl)piperidin-4-ol as a solid (559 mg).

A mixture of the solid (559 mg, 1.95 mmol) and Pd—C (10%, 90 mg) in MeOH(15 mL) containing HOAc (0.15 mL) was hydrogenated under balloon H₂overnight. It was filtered through celite. The filtrate was concentratedin vacuo to give a solid (491 mg) as1-(4-aminophenylsulfonyl)piperidin-4-ol.

To a suspension of 6-aminouracil (15.0 g, 0.118 mol) in H₂O (600 mL) wasadded NaOAc (15.0 g, 0.183 mol) and C1CH₂CHO (50% in H₂O, 30 mL). Themixture was heated at reflux for 4 h and was cooled to room temperature.The resulting dark brown precipitates were collected by filtration toafford crude 7H-pyrrolo[2,3-d]pyrimidine-2,4-diol (14.4 g, 81% yield).

A suspension of 7H-pyrrolo[2,3-d]pyrimidine-2,4-diol (2.25 g, 0.015 mol)in PhPOCl₂ was heated at 165° C. for 3 h and 180° C. for additional 3 h,the resulting dark syrup was poured slowly to ice water, the blackprecipitates were filtered off, and the filtrate was extracted withether. Ether layers were combined, washed with Sat NaHCO₃, brine, driedover Na₂SO₄ and concentrated to give2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (0.800 g, 28% yield).

To a suspension of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (0.765 g,4.05 mmol) in n-BuOH (7.5 mL) was added cyclobutyl amine (0.415 mL, 4.86mmol) and DIPEA (0.865 mL, 4.86 mmol). The mixture was heated at 65° C.for 4 h, and was then cooled to room temperature. The mixture wasdiluted with CHCl₃, washed with water, sat. NaHCO₃, brine, dried andconcentrated to give crude2-chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (0.800 g,88%), which was used directly for next step without purification.

General procedure for final coupling: a mixture of2-chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (80 mg, 0.36mmol), 1-(4-aminophenylsulfonyl)piperidin-4-ol (138 mg, 0.54 mmol) andtrimethylsilyl chloride (0.030 mL, 0.24 mmol) in nBuOH (2 mL) wasstirred at 116° C. for 4 h. The mixture was purified by reverse phaseHPLC using a gradient of 10-55% CH₃CN in water over 10 min. as eluentsto give a powder (8 mg). MS 443.1 (M+H); UV 228.5, 306.7 nm.

Example 53 butyl2-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-N-methylphenylsulfonamido)acetate

The titled compound was prepared analogously to the procedures describedin the Example for1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol.MS 487.0 (M+H); UV 206.1, 311.5 nm.

Example 542-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonamido)acetamide

The titled compound was prepared analogously to the procedures describedin the Example for1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol.MS 416.1 (M+H); UV 202.6, 227.3, 311.5 nm.

Example 552-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-N-methylphenylsulfonamido)aceticacid

To a solution of butyl2-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-N-methylphenylsulfonamido)acetate(15 mg, 0.031 mmol) in MeOH (2 mL), aqueous 1N NaOH (1.00 mL, 1.00 mmol)was added. The mixture was stirred at room temperature for 3 h. GlacialHOAc (0.20 mL) was added to neutralize NaOH. When water was added,precipitate came out, which was collected by filtration to give thetitled compound (3 mg). MS 431.0 (M+H); UV 206.1, 312.7 nm.

Example 561-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperidin-4-ol

A mixture of 1-fluoro-4-nitrobenzene (0.588 mL, 5.55 mmol),4-hydroxypiperidine (0.560 g, 5.54 mmol) and K₂CO₃ (1.50 g, 10.9 mmol)in DMF (10 mL) was stirred at room temperature overnight. Water wasadded to induce precipitation. The yellowish solid was collected byfiltration (0.90 g).

A mixture of the yellowish solid (0.90 g, 4.05 mmol) and Pd—C (10%, 120mg) in MeOH (20 mL) containing aqueous 6N HCl (0.20 mL) was hydrogenatedunder balloon H2 overnight. It was filtered through celite. The filtratewas concentrated in vacuo to give a solid (0.841 g) as1-(4-aminophenyl)piperidin-4-ol.

A mixture of 2-chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(80 mg, 0.36 mmol), 1-(4-aminophenyl)piperidin-4-ol (104 mg, 0.54 mmol)and trimethylsilyl chloride (0.030 mL, 0.24 mmol) in nBuOH (2 mL) wasstirred at 116° C. for 3 h. The mixture was purified by reverse phaseHPLC using a gradient of 5-40% CH₃CN in water over 10 min. as eluents togive a powder (5 mg). MS 379.2 (M+H); UV 204.9, 229.6, 272.2, 305.5 nm.

Example 57 tert-butyl4-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)piperidine-1-carboxylate

A mixture of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (220 mg, 1.17mmol), 4-amino-N—BOC-piperidine (280 mg, 1.40 mmol) and triethylamine(0.400 mL, 2.88 mmol) in nBuOH (8 mL) was stirred at 70° C. overnight.The mixture was purified by HPLC to give a powder (87 mg).

A mixture of the powder (40 mg, 0.11 mmol), 6-aminoindazole (40 mg, 0.30mmol) and trimethylsilyl chloride (0.020 mL, 0.16 mmol) in nBuOH (2 mL)was stirred at 116° C. overnight. The mixture was purified by reversephase HPLC using a gradient of 20-65% CH₃CN in water over 10 min. aseluents to give a powder (5 mg). MS 449.0 (M+H); UV 204.9, 242.6, 307.9nm.

Example 58

N2-(1H-indazol-6-yl)-N4-(piperidin-4-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

A solution of tert-butyl4-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)piperidine-1-carboxylate(4 mg, 0.009 mmol) in TFA (1 mL) was stirred at room temperature for 2h. It was concentrated in vacuo. The residue was purified by reversephase HPLC using a gradient of 3-30% CH₃CN in water over 10 min. aseluents to give a powder (1 mg). MS 349.0 (M+H); UV 201.4, 242.6, 306.7nm.

Example 59N4-cyclobutyl-N2-(1H-indazol-6-yl)-6-methyl-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

A mixture of 6-aminouracil) 1.00 g, 7.87 mmol), sodium acetate (1.00 g,12.2 mmol) and chloroacetone (95%, 1.00 mL, 11.9 mmol) in H2O (40 mL)was heated at reflux for 72 h. The solids were collected, and dried onvacuum to give 6-methyl-7H-pyrrolo[2,3-d]pyrimidine-2,4-diol (495 mg).

A mixture of 6-methyl-7H-pyrrolo[2,3-d]pyrimidine-2,4-diol (490 mg, 2.97mmol) in phenylphosphonic dichloride (10 mL) was heated at 180° C. for 5h. It was poured onto ice. Et2O (100 mL) was added. The organic phasewas separated, washed with 5% NaHCO₃, dried over Na₂SO₄, concentrated invacuo to give 2,4-dichloro-6-methyl-7H-pyrrolo[2,3-d]pyrimidine as asolid (380 mg).

A solution of 2,4-dichloro-6-methyl-7H-pyrrolo[2,3-d]pyrimidine (190 mg,0.94 mmol), cyclobutylamine (0.160 mL, 1.88 mmol) and triethylamine(0.200 mL, 1.44 mmol) in nBuOH (6 mL) was stirred at 70° C. overnight.It was then purified by HPLC to give2-chloro-N-cyclobutyl-6-methyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (100mg).

A mixture of2-chloro-N-cyclobutyl-6-methyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (55mg, 0.23 mmol), 6-aminoindazole (62 mg, 0.46 mmol) and trimethylsilylchloride (0.040 mL, 0.31 mmol) in nBuOH (2 mL) was heated at 116° C.overnight. It was then purified by HPLC to give the titled compound (6mg). MS 334.4 (M+H); UV 239.1, 316.2 nm

Example 60

N4-(3-amino-2,2-dimethylpropyl)-N2-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

To 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (170 mg, 0.904 mmol) insolid form in a round-bottom flask, a solution of2,2-dimethyl-1,3-propanediamine (0.325 mL, 2.71 mmol) and triethylamine(0.150 mL, 1.08 mmol) in nBuOH (4 mL) was added. The mixture was heatedat 70° C. for 3 h. It was then purified by HPLC to giveN-(3-amino-2,2-dimethylpropyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amineas a solid (212 mg).

A mixture ofN-(3-amino-2,2-dimethylpropyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amine(70 mg, 0.28 mmol), 6-aminoindazole (93 mg, 0.70 mmol) andtrimethylsilyl chloride (0.100 mL, 0.72 mmol) in nBuOH (2 mL) was heatedat 116° C. overnight. It was then purified by HPLC to give the titledcompound (15 mg). MS 351.4 (M+H); UV 206.1, 242.6, 304.3 nm.

Example 61N2-(1H-indazol-6-yl)-N4-(3-morpholinopropyl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

A solution of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (105 mg, 0.56mmol), 3-morpholinopropylamine (0.164 mL, 1.12 mmol) and triethylamine(0.150 mL, 1.08 mmol) in nBuOH (3 mL) was stirred at 60° C. overnight.It was then purified by HPLC to give2-chloro-N-(3-morpholinopropyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine as asolid (204 mg).

A mixture of2-chloro-N-(3-morpholinopropyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (65mg, 0.22 mmol), 6-aminoindazole (80 mg, 0.60 mmol) and trimethylsilylchloride (0.100 mL, 0.72 mmol) in nBuOH (3 mL) was heated at 116° C.overnight. It was then purified by HPLC to give the titled compound (16mg). MS 393.5 (M+H); UV 217.9, 239.1, 309.1 nm.

Example 62N-(4-(4-(4-aminobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A solution of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (100 mg, 0.53mmol), 1,4-diaminobutane (0.125 mL, 1.25 mmol) and triethylamine (0.150mL, 1.08 mmol) in nBuOH (4 mL) was stirred at 70° C. for 4 h. It wasthen purified by HPLC to giveN-(4-aminobutyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amine as a solid(63 mg).

A mixture ofN-(4-aminobutyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amine (63 mg,0.26 mmol), N-(4-aminophenyl)-N-methylacetamide (81 mg, 0.49 mmol) andtrimethylsilyl chloride (0.067 mL, 0.52 mmol) in nBuOH (3 mL) was heatedat 116° C. for 4 h. It was then purified by HPLC to give the titledcompound (5 mg). MS 368.2 (M+H); UV 203.8, 268.7, 301.9 nm.

Example 63N-(4-(4-(2-aminocyclohexylamino)-7H-pyrrolo[2,3-d-]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A solution of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (100 mg, 0.53mmol), cis-1,2-diaminocyclohexane (0.107 mL, 0.91 mmol) andtriethylamine (0.150 mL, 1.08 mmol) in nBuOH (4 mL) was stirred at 70°C. for 4 h. It was then purified by HPLC to giveN1-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-yl)cyclohexane-1,2-diamine asa solid (89 mg).

A mixture ofN1-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-yl)cyclohexane-1,2-diamine (44mg, 0.17 mmol), N-(4-aminophenyl)-N-methylacetamide (56 mg, 0.34 mmol)and trimethylsilyl chloride (0.146 mL, 1.15 mmol) in nBuOH (2 mL) washeated at 116° C. for 4 h. It was then purified by HPLC to give thetitled compound (6 mg). MS 394.2 (M+H); UV 201.4, 262.7, 306.7 nm.

Example 64N-(4-(4-(2-aminoethylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A solution of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (160 mg, 0.85mmol), ethylenediamine (0.170 mL, 2.55 mmol) and triethylamine (0.100mL, 0.72 mmol) in nBuOH (4 mL) was stirred at 70° C. for 4 h. It wasthen purified by HPLC to giveN-(2-aminoethyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amine as a solid(71 mg).

A mixture ofN-(2-aminoethyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amine (71 mg,0.34 mmol), N-(4-aminophenyl)-N-methylacetamide (110 mg, 0.67 mmol) andtrimethylsilyl chloride (0.085 mL, 0.67 mmol) in nBuOH (2 mL) was heatedat 116° C. for 3 h. More trimethylsilyl chloride (0.100 mL, 0.79 mmol)was added. After being stirred at 116° C. for another 2 h, the mixturewas purified by HPLC to give the titled compound (28 mg). MS 340.2(M+H); UV 201.4, 262.7, 303.1 nm.

Example 65N-(4-(4-(5-aminopentylamino)-7H-pyrrolo[2,3-d-]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A solution of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (70 mg, 0.37mmol), 1,5-diaminopentane (0.130 mL, 1.11 mmol) and triethylamine (0.150mL, 1.08 mmol) in nBuOH (3 mL) was stirred at 70° C. for 1 h. It wasthen purified by HPLC to giveN-(5-aminopentyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amine as a solid(79 mg).

A mixture ofN-(5-aminopentyl)-2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-amine (79 mg,0.31 mmol), N-(4-aminophenyl)-N-methylacetamide (100 mg, 0.61 mmol) andtrimethylsilyl chloride (0.150 mL, 1.19 mmol) in nBuOH (2 mL) was heatedat 116° C. for 4 h. The mixture was then purified by HPLC to give thetitled compound (9 mg). MS 382.2 (M+H); UV 210.8, 267.5, 305.5 nm.

Example 661-(4-(4-(4-(4-aminocyclohexylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

To a solution of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (1.12 g, 5.96mmol) in DMSO (8 mL), sodium thiomethoxide (0.500 g, 7.14 mmol) wasadded. The mixture was stirred at room temperature for 2 h. H2O wasadded to induce precipitation. The precipitate was collected and driedon vacuum to give 2-chloro-4-(methylthio)-7H-pyrrolo[2,3-d]pyrimidine asa solid (0.90 g).

A mixture of 2-chloro-4-(methylthio)-7H-pyrrolo[2,3-d]pyrimidine (484mg, 2.43 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (645 mg,2.95 mmol) and trimethylsilyl chloride (0.500 mL, 3.95 mmol) in nBuOH(10 mL) was heated at 116° C. for 48 h. After cooling down, H2O andEtOAc were added. The organic phase was separated, washed with 5%NaHCO₃, dried over Na₂SO₄, concentrated in vacuo to give1-(4-(4-(4-(methylthio)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanoneas a solid (453 mg).

To a solution of1-(4-(4-(4-(methylthio)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(451 mg, 1.18 mmol) in CH₂Cl₂ (12 mL), mCPBA (320 mg, 65%, 1.20 mmol)was added. The mixture was stirred at room temperature for 1 h. It wasthen concentrated in vacuo, the residue was purified by HPLC to give1-(4-(4-(4-(methylsulfinyl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanoneas a solid (136 mg).

To a solution of1-(4-(4-(4-(methylsulfinyl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(34 mg, 0.085 mmol) in DMSO (1 mL), 1,4-diaminocyclohexane (cis andtrans mixture, 35 mg, 0.31 mmol) and diisopropylethylamine (0.060 mL,0.35 mmol) were added. The mixture was heated at 116° C. overnight. Itwas then purified by HPLC to give the titled compound (5 mg). MS 449.3(M+H); UV 208.5, 265.1, 305.5 nm.

Example 67N-(4-(4-(5-aminopentylamino)-5-bromo-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamideandN-(4-(4-(5-aminopentylamino)-5,6-dibromo-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

To a solution ofN-(4-(4-(5-aminopentylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(44 mg, 0.12 mmol) in DMF (1 mL), N-bromosuccinamide (25 mg, 0.14 mmol)was added. The mixture was stirred at room temperature for 2 h. It wasthen purified by HPLC to give the mono-bromo compound (3 mg) and thedibromo compound (5 mg). MS 460.1 and 462.1 (mono-bromo, M+H, Brpattern); UV 207.3, 228.5, 298.4 nm; 539.0, 540.0, 541.0 and 542.0(di-bromo, M+H, 2 Br pattern); UV 202.6, 232.0, 300.7 nm.

Example 68N-(4-(4-(3-aminosulfonyl-phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A mixture of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (200 mg,0.58 mmol), 3-aminobenzenesulfonamide (111 mg, 0.65 mmol) andtriethylamine (0.200 mL, 1.44 mmol) in nBuOH (6 mL) was heated at 80° C.overnight. The mixture was then heated at 110° C. for another 24 h.After cooling down, H2O and EtOAc were added. The organic phase wasseparated, washed with 1N HCl, then with 5% NaHCO3, and it was driedover Na2SO4, concentrated in vacuo to give2-chloro-N-(3-aminosulfonyl-phenyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amineas a solid (210 mg).

A mixture of2-chloro-N-(3-aminosulfonyl-phenyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(100 mg, 0.21 mmol), N-(4-aminophenyl)-N-methylacetamide (100 mg, 0.61mmol) and trimethylsilyl chloride (0.050 mL, 0.40 mmol) in nBuOH (2 mL)and dioxane (2 mL) was heated at 110° C. for 4 h. More trimethylsilylchloride (0.100 mL, 0.79 mmol) was added. The mixture was stirred at110° C. for another 72 h. It was then purified by HPLC to giveN-(4-(4-(3-aminosulfonyl-phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamideas a solid (20 mg).

To a solution ofN-(4-(4-(3-aminosulfonyl-phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(20 mg, 0.033 mmol) in MeOH (2 mL), aq. 1N KOH (0.60 mL, 0.60 mmol) wasadded. The mixture was heated at 60° C. for 3 h. It was concentrated invacuo. The residue was acidified with HOAc (1 mL) before being purifiedby HPLC to give the titled compound (7 mg). MS 452.0 (M+H); UV 208.5,284.1, 317.4 nm.

Example 69 Ethyl2-(6-(4-(cyclobutyamino)-7H-pyrrolo-[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate

To a mixture of 6-nitro-3,4-dihydroquinolin-2(1H)-one (0.5 g, 2.6 mmol)and Cs₂CO₃ (1.70 g, 5.2 mmol) in DMF (5 mL) was added ethyl2-bromoacetate (0.317 mL, 2.86 mmol). After stirring at 65° C. for 3 h,the mixture was diluted with H₂O and EtOAc, organic layer was separated,and the aqueous layer was further extracted with EtOAc, organic layerswere combined, dried over Na₂SO₄, and was concentrated in vacuo toprovide ethyl 2-(6-nitro-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate,which was used directly for next step.

To a suspension of 1 ethyl2-(6-nitro-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate in EtOH (10 mL)and H₂O (5 mL) was added ammonium formate (1.5 g, 23.8 mmol) and Fe (0.3g, 5.4 mmol). After stirring at reflux for 2 h, the black mixture wasfiltered, the filtrate was diluted with EtOAc and water, EtOAc layer wasseparated and the aqueous layer was further extracted with EtOAc,organic layers were combined, dried over Na₂SO₄, and was concentrated invacuo to provide ethyl2-(6-amino-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate (0.2 g, 31%).

According to the general procedure for synthesis of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol,ethyl 2-(6-amino-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate and2-chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine gave Ethyl2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate(MS calcd for C₂₃H₂₆N₆O₃ 434.2, found [MH] 435.2; UV 209.8, 304.5 nm).

Example 702-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)aceticacid

To a solution of ethyl2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)acetate(10 mg, 0.023 mmol) in THF (0.6 mL) and H2O (0.4 mL) was added NaOH (5N,2 drops). After stirring for 3 h at room temperature, the reactionmixture was purified by preparative HPLC to yield2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)aceticacid (MS calcd for C₂₁H₂₂N₆O₃ 406.2, found [MH] 407.0; UV 207.5, 304.5nm).

Example 712-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)-N-(2-(dimethylamino)ethyl)acetamide

To a solution of2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)aceticacid 4 mg, 0.01 mmol) in DMF (0.3 mL) was added HATU, DIPEA andN,N-dimethylaminoethylamine. After stirring for 1 h at room temperature,it was then purified by prep HPLC to give2-(6-(4-(cyclobutyamino)-7H-pyrrolo[2,3-d]pyrimidine-2-ylamino)-2-oxo-3,4-dihydroquinolin-1(2H)-yl)-N-(2-(dimethylamino)ethyl)acetamide(MS calcd for C₂₅H₃₂N₈O₂ 476.3, found [MH] 477.0; UV 205.1, 304.5 nm).

Example 72 Methyl4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl(methyl)carbamate

According to the general procedure for synthesis of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol,methyl-4-aminophenyl(methyl)carbamate and2-Chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine gave methyl4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl(methyl)carbamate(MS calcd for C₁₉H₂₂N₆O₂ 366.2, found [MH] 368.0; UV 209.8, 271.2, 304.5nm).

Example 73 Methyl4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl(methyl)carbamate

According to the general procedure for synthesis of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol,Methyl-4-aminophenyl(methyl)carbamate and2-chloro-N-cyclopropyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine gave methyl4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl(methyl)carbamate(MS calcd for C₁₈H₂₀N₆O₂ 352.2, found [MH] 353.0; UV 227.5, 292.6 nm).

Example 74N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-hydroxy-N-methylacetamide

To a suspension of 4-nitro-N-methylaniline in DCM (10 mL) was added at0° C. was added Et₃N (0.63 mL, 4.5 mmol) and 2-benzoxyacetyl chloride(0.513 mL, 3.3 mmol). After stirring for 15 h at room temperature, thereaction mixture was diluted with EtOAc, washed with Sat. NaHCO₃, brine,dried and concentrated to give2-benzoxy-N-methyl-N-(4-nitrophenyl)acetamide (0.63 g, 70%).

To a suspension of above 2-benzoxy-N-methyl-N-(4-nitrophenyl)acetamidein EtOH (8 mL) and H₂O (4 mL) was added ammonium formate (1.3 g, 20mmol) and Fe (0.47 g, 8.4 mmol). After heating at reflux for 2 h, thereaction was cooled to room temperature, iron was filtered off, and thefiltrate was diluted with EtOAc and water, EtOAc layer was separated andthe aqueous layer was further extracted with EtOAc, organic layers werecombined, dried over Na₂SO₄, and was concentrated under vacuo to provide2-benzoxy-N-methyl-N-(4-aminophenyl)acetamide (0.4 g, 53%).

According to the general procedure for synthesis of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol,2-benzoxy-N-methyl-N-(4-aminophenyl)acetamide and2-Chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine gave2-benzyloxyN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(MS calcd for C₂₆H₂₈N₆O₂ 456.2, found [MH] 457.0).

To a solution of 2-benzyloxyN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(0.012 g, 0.026 mmol) in EtOH (0.4 mL) was added Pd/C (10 mg), and wascharged with H₂ (1 atm). After stirring for 15 h at room temperature, itwas purified by preparative HPLC to giveN-(4-(4-(cyclobutyllamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-hydroxy-N-methylacetamide(MS calcd for C₁₉H₂₂N₆O₂ 366.2, found [MH] 367.0; UV 208.7, 272.4, 304.5nm).

Example 75N-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-hydroxy-N-methylacetamide

According to the general procedure for synthesis of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol,2-benzoxy-N-methyl-N-(4-aminophenyl)acetamide and2-Chloro-N-cylclopropyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine gave2-benzyloxyN-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(MS calcd for C₂₅H₂₆N₆O₂ 442.2, found [MH] 443.5).

To a solution of 2-benzyloxyN-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(0.015 g, 0.034 mmol) in EtOH (0.4 mL) was added Pd/C (10 mg), and wascharged with H₂ (1 atm). After stirring for 15 h at room temperature, itwas purified by preparative HPLC to giveN-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-hydroxy-N-methylacetamide(MS calcd for C₁₈H₂₀N₆O₂352.2, found [MH] 352.8; UV 206.3, 228.7, 297.3nm).

Example 76N⁴-cyclobutyl-N²-(4-methylsulfonylmethyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine

To a mixture of 4-nitrobenzyl chloride (1 g, 5.8 mmol) and TBAI (0.11 g,0.29 mmol) in EtOH (12 mL) was added NaOSOCH₃ (0.77 g, 6.38 mmol). Afterheating at reflux for 2 h, the mixture was cooled to room temperature,and the precipitates were collected by filtration to give1-(methylsulfonylmethyl)-4-nitrobenzene (1.5 g).

To a mixture of 1-(methylsulfonylmethyl)-4-nitrobenzene (1.5 g, 7 mmol)in EtOH (20 mL) and H₂O (10 mL) was added iron (1.96 g, 35 mmol) andammonium formate (4.41 g, 70 mmol). After heating at 90° C. for 2 h, itwas diluted with EtOAc, washed with saturated NaHCO₃, brine, dried andconcentrated to give 4-(methylsulfonylmethyl)aniline (0.5 g).

According to the general procedure for synthesis of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol,4-(methylsulfonylmethyl)aniline and2-Chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine gaveN⁴-cyclobutyl-N²-(4-methylsulfonylmethyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine(MS calcd for C₁₈H₂₁N₅O₂S 371.5, found [MH] 372.1; UV 205.1, 273.6,304.5 nm).

Example 77N⁴-cyclobutyl-N²-(3-methylsulfonylmethyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine

To a mixture of 3-nitrobenzyl bromide (1 g, 4.63 mmol) in EtOH (12 mL)was added NaOSOCH₃ (0.61 g, 5.09 mmol). After heating at reflux for 2 h,the mixture was cooled to room temperature, and the precipitates werecollected by filtration to give 1-(methylsulfonylmethyl)-3-nitrobenzene(0.77 g).

To a mixture of 1-(methylsulfonylmethyl)-4-nitrobenzene (0.77 g, 3.56mmol) in EtOH (10 mL) and H₂O (5 mL) was added iron (1.0 g, 17.82 mmol)and ammonium formate (2.25 g, 35.6 mmol). After heating at 90° C. for 2h, it was diluted with EtOAc, washed with saturated NaHCO₃, brine, driedand concentrated to give 3-(methylsulfonylmethyl)aniline (0.47 g).

According to the general procedure for synthesis of1-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenylsulfonyl)piperidin-4-ol,3-(methylsulfonylmethyl)aniline and2-Chloro-N-cylclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine gaveN⁴-cyclobutyl-N²-(3-methylsulfonylmethyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine(MS calcd for C₁₈H₂₁N₅O₂S 371.5, found [MH] 372.1; UV 214.5, 267.6,304.5 nm).

Example 782-(3-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)phenoxy-N-methylacetamide

To a solution of 2,4-Dichloro-7H-pyrrolo[2,3-d]pyrimidine (0.062 g, 0.33mmol) in DMSO (1 mL) was added DIPEA (0.117 mL, 0.66 mmol) and2-(3-aminophenoxy)-N-methylacetamide (0.095 g, 0.49 mmol). After heatingat 100° C. for 15 h, it was diluted with EtOAc, washed with saturatedNaHCO₃, brine, dried and concentrated to give2-(3-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)phenoxy-N-methylacetamide(0.062 g).

To a mixture of2-(3-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)phenoxy-N-methylacetamide(0.062 g) in nBuOH (0.6 mL) was added 6-aminoindazole (0.028 g) andTMSCl (0.013 mL). After heating at 115° C. for 15 h, the mixture waspurified by prep HPLC to give2-(3-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)phenoxy-N-methylacetamide(0.007 g, MS calcd for C₂₂H₂₀N₈O₂ 428.4, found [MH] 429.1; UV 238.1,311.6 nm).

Example 79 N²-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine

A mixture of 2,4-Dichloro-7H-pyrrolo[2,3-d]pyrimidine (0.1 g, 0.53 mmol)in Ammonia (7 N in MeOH, 1 mL) was heated at 100° C. for 24 h, then itwas diluted with EtOAc, washed with Sat. NaHCO3, brine, dried andconcentrated to give 2-chloro-7H-pyrrolo[2,3-d]pyrimidine-4-amine (0.06g).

To a mixture of 2-chloro-7H-pyrrolo[2,3-d]pyrimidine-4-amine (0.06 g,0.353 mmol) in nBuOH (0.7 mL) was added 6-aminoindazole (0.094 g, 0.70mmol) and TMSCl (0.023 mL, 0.175 mmol). After heating at 115° C. for 8h, it was cooled and purified by prep HPLC to giveN²-(1H-indazol-6-yl)-7H-pyrrolo[2,3-d]pyrimidin-2,4-diamine (0.012 g, MScalcd for C₁₃H₁₁N₇ 265.3, found 266.1; UV 216.9, 308.0 nm).

Example 80N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide

Step 1: To a mixture of N-methyl-4-nitroaniline (2 g, 13.1 mmol) in DCM(30 mL) was added Et3N (2.75 mL, 20 mmol) and cyclopropanecarbonylchloride (1.32 mL, 14.4 mmol) at room temperature, after stirring for 1h at room temperature, the mixture was diluted with EtOAc, the organicswas washed with Sat. NaHCO₃, brine, dried and concentrated to giveN-methyl-N-(4-nitrophenyl)cyclopropanecarboxamide as crude product (3g).

To a suspension of the above crude product (3 g) in EtOH (20 mL) and H2O(10 mL) was added iron (3.67 g, 65.5 mmol) and ammonium formate (826 g,130 mml). The mixture was heated at 90° C. for 2 h, and was cooled toroom temperature. The mixture was diluted with EtOAc, washed with Sat.NaHCO₃, brine, dried and concentrated to giveN-(4-aminophenyl)-N-methylcyclopropanecarboxamide (1.6 g).

Step 2: To a suspension of2-chloro-N-cyclobutyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (67 mg, 0.3mmol) in nBuOH (0.8 mL) was addedN-(4-aminophenyl)-N-methylcyclopropanecarboxamide (85 mg, 0.45 mmol) andTMSCl (0.02 mL, 0.15 mmol). After heating at 115° C. for 2 h, themixture was cooled and purified by preparative HPLC to giveN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide(7 mg). MS found for C₂₁H₂₄N₆O as (M+H)⁺377.2. λ=208.7, 272.4, 304.5.

Example 81N-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide.MS found for C₂₀H₂₂N₆O as (M+H)⁺363.1. λ=211.0, 228.7, 296.1.

Example 82N-(4-(4-(methylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide.MS found for C₁₈H₂₀N₆O as (M+H)⁺337.1. λ=212.2, 303.3.

Example 832-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)acetamide

Step 1: To a suspension of2-chloro-4-(methylthio)-7H-pyrrolo[2,3-d]pyrimidin-4-amine (177 mg, 0.88mmol) in nBuOH (2 mL) was added TMSCl (0.111 mL, 0.88 mmol). Afterheating at 115° C. for 24 h, the mixture was cooled and purified bypreparative HPLC to give2-(2-(1H-indazol-6-ylamino)-4-(methylthio)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(53 mg).

Step 2: To a suspension of2-(2-(1H-indazol-6-ylamino)-4-(methylthio)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(53 mg, 0.18 mmol) in DCM (1 mL) was added mCPBA (50 mg, 0.187 mmol).After stirring at room temperature for 2 h, the mixture was diluted withEtOAc, washed with Sat. NaHCO₃, brine, dried and concentrated to give amixture of corresponding sulfone and sulfoxide (100 mg total).

Step 3: To a solution of the above mixture of sulfone and sulfoxide (100mg) in DMSO (1 mL) was added glycinamide hydrogen chloride (60 mg, 0.534mmol) and DIPEA (0.16 mL, 0.89 mmol). After heating at 65° C. for 48 h,the mixture was cooled and purified by preparative HPLC to give2-(2-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)acetamide(6 mg). MS found for C₁₅H₁₄N₈O as (M+H)⁺323.2. λ=244.0, 312.8.

Example 84N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-2-(dimethlamino)-N-methylacetamide

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide.MS found for C₂₁H₂₇N₇O as (M+H)⁺394.3. λ=206.3, 273.6. 304.5.

Example 85N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-N-(3-dimethylaminopropyl)benzamide

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide.MS found for C₂₂H₂₉N₇O as (M+H)⁺408.4. λ=208.7, 227.5, 312.8.

Example 86N-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-3-hydroxy-N-methypropanamide

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide.MS found for C₂₀H₂₄N₆O₂ as (M+H)⁺381.2. λ=229.8, 271.2, 304.5.

Example 87N-(4-(4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-3-hydroxy-N-methypropanamide

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN-(4-(4-(cyclobutylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylcyclopropanecarboxamide.MS found for C₁₉H₂₂N₆O₂ as (M+H)⁺367.2. λ=229.8, 296.1.

Example 88N²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

Step 1: To a suspension of2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (100 mg, 0.29 mmol) innBuOH (1 mL) was added 3-aminopyrazole (26 mg, 0.32 mmol) and DIPEA(0.057 mL, 0.32 mmol). After heating at 90° C. for 3 h, the mixture wasdiluted with EtOAc, washed with Sat. NaHCO₃, brine, dried andconcentrated to give a mixture of2-chloro-N-(1H-pyrazol-5-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-4-amine(100 mg).

Step 2: To a suspension of2-chloro-N-(1H-pyrazol-5-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-4-amine(100 mg, 0.26 mmol) in nBuOH (1 mL) was added 6-aminoindazol (68 mg,0.512 mmol) and TMSCl (0.016 mL, 0.128 mmol). After stirring at 115° C.for 15 h, the mixture was cooled and was added MeOH (1 mL), followed bya solution of KOH (100 mg) in H2O (0.5 mL). After heating at 65° C. for2 h, the solution was concentrated and the residue was purified bypreparative HPLC to giveN2-(1H-indazol-6-ylamino)-N4-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine(7 mg). MS found for C₁₆H₁₃N₉ as (M+H)⁺332.1. λ=209.8, 251.1, 310.4.

Example 89N²-(1H-indazol-6-ylamino)-N⁴-(2-methoxyethyl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamineMS found for C₁₆H₁₇N₇O as (M+H)⁺324.2. λ=246.4, 315.2.

Example 901-(4-(4-(4-(2-methoxyethylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamineMS found for C₂₂H₂₈N₆O₂ as (M+H)⁺410.2. λ=231.0, 265.3, 302.1.

Example 91N²-(1H-indazol-6-ylamino)-N⁴-(2-hydroxyethyl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamineMS found for C₁₅H₁₅N₇O as (M+H)⁺310.2. λ=241.6, 309.2.

Example 92

N⁴-(2-2-(aminoethoxy)ethyl)-N²-(1H-indazol-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamineMS found for C₁₇H₂₀N₈O as (M+H)⁺353.2. λ=241.6, 308.0.

Example 931-(4-(4-(4-(2-(2-aminoethoxy)amino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine.MS found for C₂₃H₃₁N₇O₂ as (M+H)⁺439.1. λ=266.5, 303.3.

Example 94(S)-1-(4-(4-(2-(2-aminoethoxy)ethylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)pyrrolidine-2-carboxamide

The title compound was prepared using the same synthetic schemedemonstrated in Example ofN²-(1H-indazol-6-ylamino)-N⁴-(1H-pyrazol-5-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamineMS found for C₂₂H₂₉N₇O₂ as (M+H)⁺425.1. λ=239.3, 303.3.

Example 953-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylaminobenzamide and3-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide

A solution of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (200 mg,0.580 mmol), 3-aminobenzamide (83 mg, 0.61 mmol) and triethylamine(0.200 mL, 1.44 mmol) in dioxane (5 mL) was stirred at 110° C. for 20 h.It was concentrated in vacuo. After the residue was acidified with HOAc(1 mL), it was purified by HPLC to give3-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide (85mg).

A mixture of3-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide (85mg, 0.19 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (105 mg,0.480 mmol) and trimethylsilyl chloride (0.030 mL, 0.24 mmol) in dioxane(1 mL) and nBuOH (1 mL) was stirred at 110° C. for 72 h. It wasconcentrated in vacuo. The residue was purified by HPLC to give3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide(40 mg).

To a solution of3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide(40 mg, 0.064 mmol) in MeOH (3 mL), aq. 1N KOH (1.0 mL, 1.0 mmol) wasadded. The mixture was heated at 60° C. for 3 h. It was concentrated invacuo. The residue was acidified with HOAc (1 mL) before being purifiedby HPLC to give3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide(15 mg), MS 471.1 (M+H); UV 207.3, 284.1, 311.5 nm; and3-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzamide(5 mg), MS 429.1 (M+H); UV 202.6, 279.3 nm.

Example 963-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamideand3-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamide

A mixture of2-chloro-N-(3-aminosulfonyl-phenyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(100 mg, 0.210 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (92mg, 0.42 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) innBuOH (2 mL) and dioxane (2 mL) was heated at 110° C. for 72 h. More1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (92 mg, 0.42 mmol) wasadded. The mixture was stirred at 110° C. for another 72 h. It was thenpurified by HPLC to give3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamide(40 mg)

To a solution of3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamide(40 mg, 0.061 mmol) in MeOH (3 mL), aq. 1N KOH (1.0 mL, 1.0 mmol) wasadded. The mixture was heated at 60° C. for 3 h. It was concentrated invacuo. The residue was acidified with HOAc (1 mL) before being purifiedby HPLC to give3-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamide(14 mg), MS 507.1 (M+H); UV 212.0, 282.9, 313.8 nm; and3-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzenesulfonamide(4 mg), MS 465.1 (M+H); UV 203.8, 281.7 nm.

Example 97N-(4-(4-(3-(aminomethyl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A mixture of 3-nitrobenzylamine hydrochloride (530 mg, 2.81 mmol),di-t-butyl dicarbonate (613 mg, 2.81 mmol) and TEA (0.800 mL, 5.75 mmol)in CH₂Cl₂ (8 mL) was stirred at room temperature for 72 h. It wasconcentrated in vacuo, the residue was partitioned between water andEtOAc. The organic phase was washed with 1N HCl, then with 5% NaHCO3,dried over Na2SO4, concentrated in vacuo to give tert-butyl3-nitrobenzylcarbamate as a solid (705 mg).

A mixture of tert-butyl 3-nitrobenzylcarbamate (705 mg, 2.80 mmol) andPd—C (10%, 125 mg) in EtOAc (25 mL) (containing 6 drops of HOAc) washydrogenated under balloon hydrogen for 20 h. It was filtered throughcelite. The filtrate was concentrated in vacuo to give tert-butyl3-nitrobenzylcarbamate (602 mg).

A mixture of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (200 mg,0.585 mmol), tert-butyl 3-nitrobenzylcarbamate (137 mg, 0.617 mmol) andTEA (0.200 mL, 1.44 mmol) in nBuOH (5 mL) was stirred at 110° C. for 20h. Water and EtOAc were added. The organic phase was washed with 1N HCl,then with 5% NaHCO3, dried over Na2SO4, concentrated in vacuo to givetert-butyl3-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzylcarbamate(306 mg).

A solution of tert-butyl3-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)benzylcarbamate(306 mg, 0.580 mmol), N-(4-aminophenyl)-N-methylacetamide (190 mg, 1.16mmol) and trimethylsilyl chloride (0.100 mL, 0.791 mmol) in nBuOH (5 mL)was stirred at 116° C. for 20 h. The mixture was then purified by HPLCto giveN-(4-(4-(3-(aminomethyl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(27 mg).

To a solution ofN-(4-(4-(3-(aminomethyl)phenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide(27 mg, 0.049 mmol) in MeOH (3 mL), aq. 1N KOH (1.0 mL, 1.0 mmol) wasadded. The mixture was stirred at 60° C. for 3 h. It was concentrated invacuo. The residue was acidified with HOAc (1 mL) before being purifiedby HPLC to give the titled compound (15 mg). MS 402.3 (M+H); UV 202.6,282.9, 315.0 nm.

Example 98 (R)-1-(4-(4-(4-(1hydroxypropan-2-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanoneand(R)-2-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol

A solution of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (0.35 g,1.0 mmol), D-alaminol (0.10 mL, 1.3 mmol) and TEA (0.20 mL, 1.4 mmol) indioxane (6 mL) was stirred at 70° C. for 20 h. Water and EtOAc wereadded. The organic phase was washed with 1N HCl, then with 5% NaHCO3,dried over Na2SO4, concentrated in vacuo to give(R)-2-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol(0.38 g).

A mixture of(R)-2-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol(200 mg, 0.525 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (140mg, 0.639 mmol) and trimethylsilyl chloride (0.130 mL, 1.03 mmol) innBuOH (4 mL) was stirred at 116° C. for 48 h. It was concentrated invacuo. The residue was purified by HPLC to give(R)-1-(4-(4-(4-(1-hydroxypropan-2-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(25 mg).

To a solution of(R)-1-(4-(4-(4-(1-hydroxypropan-2-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(25 mg, 0.044 mmol) in MeOH (1 mL) and dioxane (1 mL), aq. 1N KOH (1.0mL, 1.0 mmol) was added. After being stirred at 60° C. for 3 h, it wasconcentrated in vacuo. The residue was acidified with HOAc (1 mL), thenwas purified by HPLC to give(R)-1-(4-(4-(4-(1-hydroxypropan-2-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(7 mg), MS 410.5 (M+H); UV 202.6, 267.5, 303.1 nm; and(R)-2-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol(3 mg), MS 368.5 (M+H); UV 229.6, 271.0, 301.9 nm.

Example 99 (S)-1-(4-(4-(4-(1hydroxypropan-2-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanoneand(S)-2-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol

The titled compounds were synthesized analogously as(R)-1-(4-(4-(4-(1-hydroxypropan-2-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone,MS 410.5 (M+H); UV 202.0, 266.3, 301.9; and(R)-2-(2-(4-(piperazin-1-yl)phenylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)propan-1-ol,MS 368.5 (M+H); by using L-alaminol in place of D-alaminol.

Example 1001-(4-(4-(4-(1,1-dioxy-tetrahydrothiophen-3-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanoneandN2-(4-(piperazin-1-yl)phenyl)-N4-(1,1-dioxy-tetrahydrothiophen-3-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine

A mixture of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (200 mg,0.584 mmol), 3-aminotetrahydro-1H-thiophene-1,1-dione hydrochloride (100mg, 0.583 mmol) and TEA (0.325 mL, 2.34 mmol) in dioxane (5 mL) wasstirred at 70° C. for 20 h. Water and EtOAc were added. The organicphase was separated, dried over Na2SO4, concentrated in vacuo to give2-chloro-N-(1,2-dioxy-tetrahydrothiophen-3-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(208 mg).

A mixture of2-chloro-N-(1,2-dioxy-tetrahydrothiophen-3-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(208 mg, 0.472 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (165mg, 0.753 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) innBuOH (5 mL) was stirred at 120° C. for 48 h. It was concentrated invacuo. The residue was purified by HPLC to give1-(4-(4-(4-(1,1-dioxy-tetrahydrothiophen-3-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(65 mg).

To a solution of1-(4-(4-(4-(1,1-dioxy-tetrahydrothiophen-3-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(65 mg, 0.104 mmol) in MeOH (3 mL), aq. 1N KOH (1.0 mL, 1.0 mmol) wasadded. After being stirred at 60° C. for 4 h, it was concentrated invacuo. The residue was acidified with HOAc (1 mL), then was purified byHPLC to give1-(4-(4-(4-(1,1-dioxy-tetrahydrothiophen-3-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(15 mg), MS 470.5 (M+H); andN2-(4-(piperazin-1-yl)phenyl)-N4-(1,1-dioxy-tetrahydrothiophen-3-yl)-7H-pyrrolo[2,3-d]pyrimidine-2,4-diamine(3 mg), MS 428.5 (M+H).

Example 1014-(4-(cyclopropylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide

A mixture of2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(135 mg, 0.307 mmol), 4-aminobenzamide (100 mg, 0.735 mmol) andtrimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH (3 mL) wasstirred at 135° C. for 48 h. It was concentrated in vacuo. The residuewas purified by HPLC to give the titled compound (9 mg). MS 386.2 (M+H);UV 201.8, 309.8 nm.

Example 1024-(4-amino-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzoicacid

A mixture of2-chloro-N-cyclopropyl-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(245 mg, 0.557 mmol), 4-aminobenzoic acid (150 mg, 1.09 mmol) andtrimethylsilyl chloride (0.300 mL, 2.37 mmol) in nBuOH (4 mL) wasstirred at 135° C. for 24 h. It was concentrated in vacuo. The residuewas purified by HPLC to give butyl4-(4-amino-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzoate(137 mg).

To a solution of butyl4-(4-amino-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzoate(137 mg, 0.340 mmol) in THF (3 mL), aq. 1N LiOH (2.0 mL, 2.0 mmol) wasadded. The mixture was stirred at room temperature for 20 h. It wasconcentrated in vacuo. The residue was acidified with HOAc (2 mL), thenwas purified by HPLC to give the titled compound (55 mg). MS 347.2(M+H); UV 203.8, 306.7 nm.

Example 1034-(cyclopropylamino)-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

A mixture of2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(145 mg, 0.374 mmol), 6-amino-3,4-dihydroquinolin-2(1H)-one (60 mg,0.370 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH(4 mL) was stirred at 135° C. for 20 h. It was concentrated in vacuo.The residue was purified by HPLC to give the titled compound (50 mg). MS360.3 (M+H); UV 202.2, 292.7 nm.

Example 1044-(5-cyano-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide

A mixture of2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(175 mg, 0.451 mmol), 4-aminobenzamide (122 mg, 0.897 mmol) andtrimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH (4 mL) wasstirred at 135° C. for 20 h. It was concentrated in vacuo. The residuewas purified by HPLC to give the titled compound (43 mg). MS 334.3(M+H); UV 204.6, 230.2, 308.1 nm.

Example 105N-(4-(5-cyano-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)phenyl)-N-methylacetamide

A mixture of2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(165 mg, 0.425 mmol), N-(4-aminophenyl)-N-methylacetamide (107 mg, 0.652mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH (4 mL)was stirred at 135° C. for 20 h. It was concentrated in vacuo. Theresidue was purified by HPLC to give the titled compound (50 mg). MS362.3 (M+H); UV 202.8, 254.0, 280.4 nm.

Example 1064-(5-cyano-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzenesulfonamide

A mixture of2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(150 mg, 0.387 mmol), 4-aminobenzenesulfonamide (103 mg, 0.599 mmol) andtrimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH (4 mL) wasstirred at 135° C. for 20 h. It was concentrated in vacuo. The residuewas purified by HPLC to give the titled compound (45 mg). MS 370.3(M+H); UV 202.8, 254.6, 305.0 nm.

Example 1072-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(cyclopropylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

A mixture of2-chloro-4-(cyclopropylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(150 mg, 0.387 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (132mg, 0.602 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) innBuOH (4 mL) was stirred at 135° C. for 20 h. It was concentrated invacuo. The residue was purified by HPLC to give the titled compound (25mg). MS 417.4 (M+H); UV 203.4, 254.0, 291.5 nm.

Example 1084-(cyclopropylamino)-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide

A solution of4-(cyclopropylamino)-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(30 mg, 0.083 mmol), K₂CO₃ (100 mg, 0.724 mmol) and H₂O₂ (50% aq., 0.500mL) in DMSO (1 mL) was heated at 100° C. for 5 min. Gas evolvedviolently. After cooling down, the mixture was purified by HPLC to givethe titled compound (7 mg). MS 378.4 (M+H); UV 200.4, 241.8, 295.8 nm.

Example 1094-(4-(1-(2-cyanoacetyl)piperidin-4-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide

A mixture of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (171 mg,0.500 mmol), 4-amino-1-N-Boc-piperidine (100 mg, 0.500 mmol) and TEA(0.150 mL, 1.08 mmol) in dioxane (4 mL) was stirred at 70° C. for 20 h.Water and EtOAc were added. The organic phase was separated, washed with1N HCl, then with 5% NaHCO3, dried over Na2SO4, concentrated in vacuo togive tert-butyl4-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)piperidine-1-carboxylate(202 mg).

A mixture of tert-butyl4-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)piperidine-1-carboxylate(202 mg, 0.400 mmol), 4-aminobenzamide (82 mg, 0.60 mmol) andtrimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH (4 mL) wasstirred at 116° C. in a sealed tube for 20 h. It was concentrated invacuo. The residue was purified by HPLC to give4-(4-(piperidin-4-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(58 mg).

To a suspension of cyanoacetic acid (85 mg, 1.00 mmol) in CH₂Cl₂ (2 mL)(containing 2 drops of DMF), oxalyl chloride (0.082 mL, 0.94 mmol) wasadded. The mixture was stirred at room temperature for 50 min. To theabove solution cooled in an ice bath, a solution of4-(4-(piperidin-4-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(58 mg, 0.11 mmol) and TEA (0.450 mL, 3.22 mmol) in DMF (2 mL) wasadded. The mixture was stirred for 2 h. Water and EtOAc were added. Theorganic phase was separated, washed with 5% NaHCO3, dried over Na2SO4,concentrated in vacuo to give4-(4-(1-(2-cyanoacetyl)piperidin-4-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(45 mg).

To a solution of4-(4-(1-(2-cyanoacetyl)piperidin-4-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(45 mg, 0.078 mmol) in dioxane (2 mL) and MeOH (1 mL), aq. 1N KOH (1.0mL, 1.0 mmol) was added. The mixture was stirred at 70° C. for 4 h. Itwas concentrated in vacuo. The residue was acidified with HOAc (1 mL)before being purified by HPLC to give4-(4-(piperidin-4-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(20 mg).

To a suspension of cyanoacetic acid (85 mg, 1.00 mmol) in CH₂Cl₂ (2 mL)(containing 2 drops of DMF), oxalyl chloride (0.082 mL, 0.94 mmol) wasadded. The mixture was stirred at room temperature for 50 min. To theabove solution cooled in an ice bath, a solution of4-(4-(piperidin-4-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(20 mg, 0.057 mmol) and TEA (0.450 mL, 3.22 mmol) in DMF (2 mL) wasadded. The mixture was stirred at room temperature for 20 h. Water wasadded. CH₂Cl₂ was removed in vacuo. The residue was purified by HPLC togive the titled compound (10 mg). MS 419.3 (M+H); UV 200.0, 311.9 nm.

Example 1104-(4-(1-(2-cyanoacetyl)piperidin-3-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide

A mixture of 2,4-dichloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine (233 mg,0.681 mmol), 3-amino-1-N-Boc-piperidine (136 mg, 0.680 mmol) and TEA(0.200 mL, 1.44 mmol) in dioxane (5 mL) was stirred at 70° C. for 20 h.Water and EtOAc were added. The organic phase was separated, washed with1N HCl, then with 5% NaHCO3, dried over Na2SO4, concentrated in vacuo togive tert-butyl3-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)piperidine-1-carboxylate(251 mg).

A mixture of tert-butyl3-(2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamino)piperidine-1-carboxylate(251 mg, 0.496 mmol), 4-aminobenzamide (102 mg, 0.750 mmol) andtrimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH (4 mL) wasstirred at 116° C. in a sealed tube for 20 h. It was concentrated invacuo. The residue was purified by HPLC to give4-(4-(piperidin-3-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(36 mg).

To a solution of4-(4-(piperidin-3-ylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(36 mg, 0.071 mmol) in dioxane (2 mL) and MeOH (2 mL), aq. 1N KOH (1.0mL, 1.0 mmol) was added. The mixture was stirred at 70° C. for 3 h. Itwas concentrated in vacuo. The residue was acidified with HOAc (1 mL)before being purified by HPLC to give the desired compound as TFA salt,which was then dissolved in MeOH (5 mL), MP-carbonate (0.10 g, 0.31mmol) was added. After gentle agitation for 30 min, the mixture wasfiltered. The filtrate was concentrated in vacuo to give4-(4-(piperidin-3-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(18 mg) as free base.

To a suspension of cyanoacetic acid (46 mg, 0.54 mmol) in CH₂Cl₂ (1 mL)(containing 2 drops of DMF), oxalyl chloride (0.047 mL, 0.54 mmol) wasadded. The mixture was stirred at room temperature for 40 min. To theabove solution cooled in an ice bath, a solution of4-(4-(piperidin-3-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)benzamide(18 mg, 0.051 mmol) and TEA (0.250 mL, 1.80 mmol) in DMF (2 mL) wasadded. The mixture was stirred for 2 h. Water was added. CH₂Cl₂ wasremoved in vacuo. The residue was purified by HPLC to give the titledcompound (10 mg). MS 419.3 (M+H); UV 201.6, 311.9 nm.

Example 1114-(4-methoxyphenylamino)-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

A mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(260 mg, 0.556 mmol), p-anisidine (75 mg, 0.610 mmol) and DIEA (0.300mL, 1.72 mmol) in dioxane (5 mL) was stirred at 110° C. for 20 h. Waterand EtOAc were added. The organic phase was separated, washed with 1NHCl, then with 5% NaHCO3, dried over Na2SO4, concentrated in vacuo togive2-chloro-5-iodo-N-(4-methoxyphenyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(257 mg).

A solution of2-chloro-5-iodo-N-(4-methoxyphenyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(257 mg, 0.463 mmol), Pd2 dba3 (25 mg, 0.027 mmol) and dppf (51 mg,0.092 mmol) in DMF (4 mL) was degassed with argon before being chargedwith Zn(CN)2 (65 mg, 0.555 mmol). The mixture was stirred at 70° C. for3 h. DMF was removed in vacuo. The residue was dissolved in CH₃CN. Waterwas added to induce precipitation. The precipitate was collected anddried on vacuum to give2-chloro-4-(4-methoxyphenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(68 mg).

A mixture of2-chloro-4-(4-methoxyphenylamino)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(68 mg, 0.15 mmol), 6-amino-3,4-dihydroquinolin-2(1H)-one (40 mg, 0.24mmol) and trimethylsilyl chloride (0.100 mL, 0.790 mmol) in nBuOH (2 mL)was stirred at 135° C. for 20 h. More6-amino-3,4-dihydroquinolin-2(1H)-one (24 mg, 0.15 mmol) andtrimethylsilyl chloride (0.200 mL, 1.58 mmol) were added. It was stirredat 135° C. for another 20 h. nBuOH was removed in vacuo. The residue waspurified by HPLC to give the titled compound (5 mg). MS 426.4 (M+H); UV203.5, 290.3 nm.

Example 1126-(4-(4-fluorophenylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-1-methyl-3,4-dihydroquinolin-2(1H)-one

A mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(260 mg, 0.556 mmol), 4-fluoroaniline (0.055 mL, 0.581 mmol) and DIEA(0.300 mL, 1.72 mmol) in dioxane (4 mL) was stirred at 110° C. for 20 h.Water and EtOAc were added. The organic phase was separated, washed with1N HCl, then with 5% NaHCO3, dried over Na2SO4, concentrated in vacuo togive2-chloro-N-(4-fluorophenyl)-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(230 mg).

To a mixture of2-chloro-N-(4-fluorophenyl)-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(230 mg, 0.423 mmol), pyridine-4-boronic acid (57 mg, 0.463 mmol) andPd(Ph3P)2Cl2 (30 mg, 0.042 mmol) in dioxane (4 mL), aq. Na2CO3 (135 mg,1.27 mmol) (1.0 mL) was added. The mixture was stirred at 100° C. for 2h. It was concentrated in vacuo. The residue was triturated. The solidwas collected by filtration (86 mg). The filtrate was purified by HPLCto give2-chloro-N-(4-fluorophenyl)-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(31 mg).

A solution of2-chloro-N-(4-fluorophenyl)-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(31 mg, 0.062 mmol), 6-amino-1-methyl-3,4-dihydroquinolin-2(1H)-one (25mg, 0.14 mmol) and trimethylsilyl chloride (0.100 mL, 0.790 mmol) innBuOH (1 mL) was stirred at 135° C. for 20 h. It was concentrated invacuo. The residue was purified by HPLC to give the titled compound. MS480.5 (M+H); UV 204.7, 281.7, 314.3 nm.

Example 1136-(4-Amino-5-(4-fluorophenyl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-1-methyl-3,4-dihydroquinolin-2(1H)-one

To a mixture of2-chloro-N-cyclopropyl-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(150 mg, 0.307 mmol), 4-fluorophenylboronic acid (64 mg, 0.457 mmol) andPd(Ph3P)2Cl2 (22 mg, 0.031 mmol) in dioxane (3 mL), aq. Na2CO3 (100 mg,0.943 mmol) (1.0 mL) was added. The mixture was stirred at 100° C. for 1h. Water and EtOAc were added. The organic phase was separated, driedover Na2SO4, concentrated in vacuo. The residue was purified by a silicagel column, which was eluted with 5-15% EtOAc in hexane to give2-chloro-N-cyclopropyl-5-(4-fluorophenyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(121 mg).

A solution of2-chloro-N-cyclopropyl-5-(4-fluorophenyl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(76 mg, 0.17 mmol), 6-amino-1-methyl-3,4-dihydroquinolin-2(1H)-one (60mg, 0.34 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) innBuOH (2 mL) was stirred at 135° C. for 20 h. It was concentrated invacuo. The residue was purified by HPLC to give the titled compound. MS403.4 (M+H); UV 205.3, 293.4 nm.

Example 1146-(4-(benzylamino)-5-(pyridin-4-yl)-7H-pyrrolo[2,3-d]pyrimidin-2-ylamino)-1-methyl-3,4-dihydroquinolin-2(1H)-one

A mixture of 2,4-dichloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine(1.00 g, 2.13 mmol), benzylamine (0.233 mL, 2.13 mmol) and DIEA (0.750mL, 4.31 mmol) in dioxane (12 mL)/nBuOH (12 mL)/DMF (3 mL) was stirredat 70° C. for 2 h. It was concentrated in vacuo. The residue waspartitioned between water and EtOAc. The organic phase was separated,washed with 1N HCl, then with 5% NaHCO3, dried over Na2SO4, concentratedin vacuo to giveN-benzyl-2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(0.993 g).

To a mixture ofN-benzyl-2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (300mg, 0.557 mmol), pyridine-4-boronic acid (137 mg, 1.11 mmol) andPd(Ph3P)₂C12 (40 mg, 0.042 mmol) in dioxane (3 mL), aq. Na2CO3 (120 mg,1.13 mmol) (1.0 mL) was added. The mixture was stirred at 100° C. for 4h. It was concentrated in vacuo. The residue was purified by HPLC togiveN-benzyl-2-chloro-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(27 mg).

A solution ofN-benzyl-2-chloro-5-(pyridin-4-yl)-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine(27 mg, 0.055 mmol), 6-amino-1-methyl-3,4-dihydroquinolin-2(1H)-one (20mg, 0.11 mmol) and trimethylsilyl chloride (0.070 mL, 0.553 mmol) innBuOH (1.5 mL) was stirred at 116° C. for 20 h. More trimethylsilylchloride (0.100 mL, 0.790 mmol) was added. It was stirred at 116° C. foranother 40 h. It was concentrated in vacuo. The residue was purified byHPLC to give the titled compound. MS 476.4 (M+H); UV 200.3, 268.7, 309.3nm.

Example 1154-(Benzylamino)-2-(1-methyl-2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile

A solution ofN-benzyl-2-chloro-5-iodo-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-4-amine (386mg, 0.716 mmol), Pd2 dba3 (40 mg, 0.043 mmol) and dppf (80 mg, 0.14mmol) in DMF (6 mL) was degassed with argon before being charged withZn(CN)2 (100 mg, 0.555 mmol). The mixture was stirred at 70° C. for 20h. DMF was removed in vacuo. The residue was purified by HPLC to give4-(benzylamino)-2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(54 mg).

A solution of4-(benzylamino)-2-chloro-7-tosyl-7H-pyrrolo[2,3-d]pyrimidine-5-carbonitrile(54 mg, 0.12 mmol), 6-amino-1-methyl-3,4-dihydroquinolin-2(1H)-one (43mg, 0.24 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) innBuOH (2.5 mL) and dioxane (1.5 mL) was stirred at 116° C. for 20 h. Itwas concentrated in vacuo. The residue was purified by HPLC to give thetitled compound. MS 424.3 (M+H); UV 207.7, 273.1 nm.

Example 116

Butyl1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylate

A solution of 2,6-dichloropurine (189 mg, 1.00 mmol), ethyl nipecotate(0.155 mL, 1.00 mmol) and DIEA (0.300 mL, 1.72 mmol) in CH₃CN (5 mL) wasstirred at room temperature for 20 h, during which time white solidsprecipitated out, which was collected to give ethyl1-(2-chloro-9H-purin-6-yl)piperidine-3-carboxylate (108 mg).

A mixture of ethyl 1-(2-chloro-9H-purin-6-yl)piperidine-3-carboxylate(108 mg, 0.349 mmol), 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone (115mg, 0.525 mmol) and trimethylsilyl chloride (0.300 mL, 2.37 mmol) innBuOH (4 mL) was stirred at 135° C. for 20 h. It was concentrated invacuo. The residue was purified by HPLC to give the titled compound (86mg). MS 521.4 (M+H); UV 201.8, 272.8 nm.

Example 1171-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylicacid and1-(2-(4-(piperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylicacid

To a solution of butyl1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylate(76 mg, 0.15 mmol) in THF (2 mL), aq. 1N LiOH (1.00 mL, 1.00 mmol) wasadded. The mixture was stirred at room temperature for 68 h. HOAc (1 mL)was added to neutralize LiOH. The mixture was then purified by HPLC togive1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylicacid (45 mg), MS 465.3 (M+H); UV 205.8, 272.8 nm; and1-(2-(4-(piperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylicacid (8 mg), MS 423.3 (M+H).

Example 1181-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxamide

To a solution of1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-9H-purin-6-yl)piperidine-3-carboxylicacid (40 mg, 0.086 mmol) and HOBt (30 mg, 0.196 mmol) in DMF (2 mL), EDC(35 mg, 0.183 mmol) was added. After being stirred for 90 min, NH3 (0.5M in dioxane, 1.00 mL, 0.500 mmol) was added. The mixture was stirredfor 2 h. More EDC (52 mg, 0.271 mmol) was added. It was stirred foranother 20 h. The mixture was then purified by HPLC to give the titledcompound (30 mg). MS 464.3 (M+H); UV 204.8, 273.8 nm.

Example 119

Butyl2-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)acetate

A solution of 2,4-dichloro-7H-pyrrolo[2,3-d]pyrimidine (188 mg, 1.00mmol), 2-(4-piperidinyl)acetic acid ethyl ester (171 mg, 1.00 mmol) andDIEA (0.400 mL, 2.30 mmol) in dioxane (6 mL) was stirred at 70° C. for20 h. Water and EtOAc were added. The organic phase was separated,washed with 1N HCl, dried over Na2SO4, concentrated in vacuo to giveethyl2-(1-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)acetate(308 mg).

A mixture of ethyl2-(1-(2-chloro-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)acetate(135 mg, 0.419 mmol), 6-amino-3,4-dihydroquinolin-2(1H)-one (97 mg,0.600 mmol) and trimethylsilyl chloride (0.200 mL, 1.58 mmol) in nBuOH(4 mL) was stirred at 135° C. for 20 h. More trimethylsilyl chloride(0.200 mL, 1.58 mmol) was added. It was stirred at 135° C. for another20 h. It was concentrated in vacuo. The residue was purified by HPLC togive the titled compound (54 mg). MS 477.3 (M+H); UV 200.0, 279.8 nm.

Example 1202-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)aceticacid

To a solution of butyl2-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)acetate(48 mg, 0.101 mmol) in THF (2 mL), aq. 1N LiOH (1.00 mL, 1.00 mmol) wasadded. The mixture was stirred at room temperature for 20 h. HOAc (1 mL)was added to neutralize LiOH. It was then purified by HPLC to give thetitled compound (32 mg). MS 421.3 (M+H); UV 207.8, 279.8 nm

Example 1212-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)acetamide

To a solution of2-(1-(2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperidin-4-yl)aceticacid (26 mg, 0.062 mmol) and HOBt (35 mg, 0.23 mmol) in DMF (2 mL), EDC(40 mg, 0.21 mmol) was added. After being stirred for 40 min, NH3 (0.5 Min dioxane, 1.00 mL, 0.500 mmol) was added. The mixture was stirred for20 h. It was then purified by HPLC to give the titled compound (20 mg).MS 420.3 (M+H); UV 201.8, 279.8 nm.

Example 1221-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-chloropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

The titled compound was synthesized analogously as compound1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone,by using 2,4,5-trichloropyrimidine. MS 444.3 and 446.3 (M+H, Clpattern); UV 210.7, 265.7 nm.

Example 1236-(5-fluoro-4-(3-oxopiperazin-1-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

The titled compound was synthesized analogously as compound1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone,by using piperazin-2-one. MS 414.3 (M+H); UV 214.8, 252.8 nm.

Example 1241-(4-(4-(5-fluoro-4-(piperazin-1-yl)pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

The titled compound was synthesized analogously as compound1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone,by using 1-Boc-piperazine. MS 400.3 (M+H); UV 203.8, 268.8 nm.

Example 1254-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperazine-1-carboxamide

To a suspension of1-(4-(4-(5-fluoro-4-(piperazin-1-yl)pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(53 mg, 0.13 mmol) in CH₃CN (2 mL), a solution of KCN (60 mg, 0.74 mmol)in H2O (2 mL) was added. The suspension became clear. The mixture wasstirred at 70° C. for 3 h. It was then purified by HPLC to give thetitled compound (15 mg). MS 443.3 (M+H); UV 201.8, 269.8 nm.

Example 1262-(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)acetamide

The titled compound was synthesized analogously as compound1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone,by using 2-(piperidin-4-yl)acetamide. MS 456.3 (M+H); UV 203.8, 269.8nm.

Example 127 Benzyl1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-ylcarbamate

The titled compound was synthesized analogously as compound1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone,by using benzyl piperidin-4-ylcarbamate. MS 548.4 (M+H); UV 215.8,258.8, 279.8 nm.

Example 1281-(4-(4-(4-(4-aminopiperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

A solution of benzyl1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-ylcarbamate(370 mg, 0.676 mmol) and Pd—C (10%, 55 mg) in MeOH (12 mL) washydrogenated under balloon hydrogen for 20 h. It was then filteredthrough celite. The filtrate was concentrated in vacuo to give thetitled compound (239 mg). MS 414.3 (M+H); UV 202.8, 268.8 nm.

Example 1291-(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)urea

To a suspension of1-(4-(4-(4-(4-aminopiperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(48 mg, 0.12 mmol) in CH₃CN (2 mL), a solution of KCN (60 mg, 0.74 mmol)in H2O (2 mL) was added. The suspension became clear. The mixture wasstirred at 70° C. for 2 h. It was then purified by HPLC to give thetitled compound (12 mg). MS 457.3 (M+H); UV 204.8, 266.8 nm.

Example 1302-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(ureidomethyl)piperidin-1-yl)pyrimidine-5-carboxamide

To a suspension of2-(4-(4-acetylpiperazin-1-yl)phenylamino)-4-(4-(aminomethyl)piperidin-1-yl)pyrimidine-5-carboxamide(8 mg, 0.018 mmol) in CH₃CN (1 mL), a solution of KCN (18 mg, 0.22 mmol)in H2O (1 mL) was added. The mixture was stirred at 70° C. for 20 h. Itwas then purified by HPLC to give the titled compound (6 mg). MS 496.5(M+H); UV 200.8, 271.8 nm.

Example 1314-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzoicacid

The titled compound was synthesized analogously as compound1-(4-(4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone,by using 4-aminobenzoic acid. MS 346.2 (M+H); UV 215.8, 275.8, 292.8 nm.

Example 1326-(4-(4-(aminomethyl)piperidin-1-yl)-5-bromopyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

The titled compound was synthesized analogously as compound6-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one,by using 5-bromo-2,4-dichloropyrimidine. MS 431.4 and 433.3 (M+H, Brpattern); UV 222.3, 263.9 nm.

Example 1334-(4-(4-((dimethylamino)methyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzamide

To a mixture of4-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)benzamide(100 mg, 0.291 mmol) and HCHO (37% aq., 0.060 mL, 0.807 mmol) in MeOH (3mL) and HOAc (0.3 mL), NaBH3CN (60 mg, 0.952 mmol) was added. Themixture was stirred at room temperature for 20 h. It was concentrated invacuo. The residue was purified by HPLC to give the titled compound (20mg). MS 373.3 (M+H); UV 213.8, 269.8, 292.8 nm.

Example 134 Ethyl3-amino-3-(1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)propanoate

To a suspension of ethyl 3-(1-Boc-piperidine-4-yl)-beta-DL-alaninatehydrochloride (518 mg, 1.54 mmol) in CH₂Cl₂ (10 mL), a solution ofNaHCO3 (1.00 g, 1.20 mmol) in H2O (15 mL) was added. Benzylchloroformate (0.246 mL, 95%, 1.66 mmol) was then added. The mixture wasstirred at room temperature for 20 h. The CH₂Cl₂ layer was separated,dried over Na2SO4, concentrated in vacuo to give tert-butyl4-(1-(benzyloxycarbonylamino)-3-ethoxy-3-oxopropyl)piperidine-1-carboxylate(660 mg).

To a solution of tert-butyl4-(1-(benzyloxycarbonylamino)-3-ethoxy-3-oxopropyl)piperidine-1-carboxylate(660 mg, 1.52 mmol) in CH₂Cl₂ (5 mL), TFA (5 mL) was added. The mixturewas stirred at room temperature for 3 h. It was concentrated in vacuo.The residue was purified by HPLC to give ethyl3-(benzyloxycarbonylamino)-3-(piperidin-4-yl)propanoate as TFA salt. Thesalt was dissolved in EtOAc (50 mL), which was washed with 5% NaHCO3,dried over Na2SO4, concentrated in vacuo to give ethyl3-(benzyloxycarbonylamino)-3-(piperidin-4-yl)propanoate as free base(102 mg).

To a solution of ethyl3-(benzyloxycarbonylamino)-3-(piperidin-4-yl)propanoate (102 mg, 0.305mmol) and 2,4-dichloro-5-fluoropyrimidine (51 mg, 0.305 mmol) in CH₃CN(2 mL), DIEA (0.106 mL, 0.610 mmol) was added. The mixture was stirredat room temperature for 68 h. It was then concentrated in vacuo. Theresidue was dissolved in nBuOH (3 mL),6-amino-3,4-dihydroquinolin-2(1H)-one (82 mg, 0.506 mmol) was added. Thesolution was stirred at 116° C. for 20 h. It was concentrated in vacuo.The residue was dissolved in MeOH (9 mL). To the solution, Pd—C (10%, 40mg) was added. The mixture was then hydrogenated under balloon hydrogenfor 20 h. It was filtered through celite. The filtrate was concentratedin vacuo. The residue was purified by HPLC to give the titled compound(66 mg). MS 457.4 (M+H); UV 211.9, 277.3 nm.

Example 135 Methyl2-amino-2-(1-(5-fluoro-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-yl)piperidin-4-yl)acetate

The titled compound was synthesized analogously as compound6-(4-(4-(aminomethyl)piperidin-1-yl)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one,by using methyl 2-(tert-butoxycarbonylamino)-2-(piperidin-4-yl)acetate.MS 429.3 (M+H); UV 207.0, 274.2 nm

Example 136N-(1-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperidin-4-yl)-2-cyanoacetamide

To a suspension of cyanoacetic acid (85 mg, 1.00 mmol) in CH₂Cl₂ (2 mL)(containing 2 drops of DMF) at room temperature, oxalyl chloride (0.082mL, 0.94 mmol) was added. Gas evolved and suspension became clear. Itwas then stirred for 30 min. To the above solution cooled in an icebath, a solution of1-(4-(4-(4-(4-aminopiperidin-1-yl)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(70 mg, 0.17 mmol) and TEA (0.478 mL, 3.43 mmol) in DMF (2 mL) was addeddrop wise. The mixture was then stirred at room temperature for 3 h. Itwas concentrated in vacuo. The residue was purified by HPLC to give thetitled compound (12 mg). MS 481.5 (M+H); UV 204.7, 266.3 nm

Example 1373-(4-(2-(4-(4-acetylpiperazin-1-yl)phenylamino)-5-fluoropyrimidin-4-yl)piperazin-1-yl)-3-oxopropanenitrile

To a suspension of cyanoacetic acid (85 mg, 1.00 mmol) in CH₂Cl₂ (2 mL)(containing 2 drops of DMF) at room temperature, oxalyl chloride (0.082mL, 0.94 mmol) was added. Gas evolved and suspension became clear. Itwas then stirred for 50 min. To the above solution cooled in an icebath, a solution of1-(4-(4-(5-fluoro-4-(piperazin-1-yl)pyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone(23 mg, 0.057 mmol) and TEA (0.400 mL, 2.87 mmol) in DMF (1 mL) wasadded drop wise. The mixture was then stirred at room temperature for 90min. It was concentrated in vacuo. The residue was purified by HPLC togive the titled compound (17 mg). MS 467.5 (M+H); UV 201.0, 269.4 nm

Example 1386-(4-(4-(aminomethyl)piperidin-1-yl)-5-(pyridin-4-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

To a mixture of 5-bromo-2,4-dichloropyrimidine (0.256 mL, 2.00 mmol) and4-N-Boc-aminomethylpiperidine (428 mg, 2.00 mmol) in CH₃CN (5 mL), DIEA(0.700 mL, 4.02 mmol) was added. The mixture was stirred at roomtemperature for 40 h. Water and EtOAc were added. The organic phase wasseparated, washed with 1N HCl, then with 5% NaHCO3, dried over Na2SO4,concentrated in vacuo to give tert-butyl(1-(5-bromo-2-chloropyrimidin-4-yl)piperidin-4-yl)methylcarbamate (610mg).

To a mixture of tert-butyl(1-(5-bromo-2-chloropyrimidin-4-yl)piperidin-4-yl)methylcarbamate (229mg, 0.564 mmol), pyridine-4-boronic acid (76 mg, 0.618 mmol) andPd(Ph₃P)₂Cl₂ (40 mg, 0.056 mmol) in dioxane (3 mL), aq. Na₂CO₃ (180 mg,1.69 mmol) (1.0 mL) was added. The mixture was stirred at 100° C. for 20h. It was concentrated in vacuo. The residue was purified by HPLC togive tert-butyl(1-(2-chloro-5-(pyridin-4-yl)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(80 mg).

A solution of tert-butyl(1-(2-chloro-5-(pyridin-4-yl)pyrimidin-4-yl)piperidin-4-yl)methylcarbamate(80 mg, 0.20 mmol) and 6-amino-3,4-dihydroquinolin-2(1H)-one (50 mg,0.30 mmol) in nBuOH (3 mL) was stirred at 116° C. for 20 h. It wasconcentrated in vacuo. The residue was dissolved in TFA (4 mL). Afterbeing stirred at room temperature for 30 min, TFA was removed in vacuo.The residue was purified by HPLC to give the titled compound. MS 430.4(M+H); UV 207.7, 264.5 nm.

Example 1396-(4-(4-(aminomethyl)piperidin-1-yl)-5-cyclopropylpyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

A mixture of tert-butyl(1-(5-bromo-2-chloropyrimidin-4-yl)piperidin-4-yl)methylcarbamate (112mg, 0.276 mmol), cyclopropylboronic acid (40 mg, 0.465 mmol), K3PO4 (200mg, 0.943 mmol) and tricyclohexyl phosphine (20 mg, 0.071 mmol) intoluene (3 mL) and H2O (0.2 mL) was degassed with argon, then Pd(OAc)₂(10 mg, 0.044 mmol) was added. The mixture was stirred at 100° C. for 20h. Water and EtOAc were added. The organic phase was separated, washedwith brine, dried over Na2SO4, concentrated in vacuo to give tert-butyl(1-(2-chloro-5-cyclopropylpyrimidin-4-yl)piperidin-4-yl)methylcarbamate(130 mg).

A solution of tert-butyl(1-(2-chloro-5-cyclopropylpyrimidin-4-yl)piperidin-4-yl)methylcarbamate(99 mg, 0.27 mmol) and 6-amino-3,4-dihydroquinolin-2(1H)-one (50 mg,0.30 mmol) in nBuOH (3 mL) was stirred at 116° C. for 20 h. It was thenstirred at 140° C. for another 20 h. It was concentrated in vacuo. Theresidue was purified by HPLC to give a powder, which was then dissolvedin TFA (3 mL). After being stirred at room temperature for 20 h, TFA wasremoved in vacuo. The residue was purified by HPLC to give the titledcompound (16 mg). MS 393.5 (M+H); UV 208.9, 264.5 nm

Example 1406-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

To a mixture of trans-4-aminocyclohexanol (2.07 g, 13.6 mmol) and NaHCO3(3.50 g, 41.7 mmol) in H2O (20 mL) at room temperature, a solution ofbenzyl chloroformate (1.92 mL, 13.6 mmol) in dioxane (15 mL) was added.The mixture was stirred at room temperature for 20 h. The whiteprecipitate was collected as benzyl (1R,4R)-4-hydroxycyclohexylcarbamate(3.37 g).

To a suspension of benzyl (1R,4R)-4-hydroxycyclohexylcarbamate (1.14 g,4.58 mmol) and triethylamine (1.30 mL, 9.34 mmol) in CH₂Cl₂ (15 mL) atroom temperature, methanesulfonyl chloride (0.425 mL, 5.49 mmol) wasadded. The mixture was stirred at room temperature for 20 h. Moremethanesulfonyl chloride (0.425 mL, 5.49 mmol) and triethylamine (1.00mL) were added. Stirring was continued for 48 h. The reaction solutionwas washed with 5% NaHCO₃, then with 1 N HCl. The organic phase wasseparated, dried over Na₂SO₄, concentrated in vacuo to give(1R,4R)-4-(benzyloxycarbonyl)cyclohexyl methanesulfonate as a solid(1.13 g).

A mixture of (1R,4R)-4-(benzyloxycarbonyl)cyclohexyl methanesulfonate(1.13 g, 3.46 mmol) and NaN₃ (0.674 g, 10.4 mmol) in DMF (10 mL) wasstirred at 100° C. for 20 h. Water and EtOAc were added. The organicphase was separated, washed with water, dried over Na₂SO₄, concentratedin vacuo to give benzyl (1s,4s)-4-azidocyclohexylcarbamate (0.819 g).

To a solution of benzyl (1s,4s)-4-azidocyclohexylcarbamate (0.410 g,1.50 mmol) in THF (8 mL) and H₂O (0.100 mL, 5.56 mmol) at roomtemperature, Ph₃P (0.590 g, 2.25 mmol) was added. The solution wasstirred at 70° C. for 20 h. EtOAc and 1N HCl were added. The aqueousphase was separated, washed with EtOAc. It was then basified with 5NNaOH to pH 12. The free amine product was extracted with EtOAc. TheEtOAc solution was dried over Na₂SO₄, and concentrated in vacuo to givebenzyl (1s,4s)-4-aminocyclohexylcarbamate (0.270 g).

To a mixture of 2,4-dichloro-5-fluoropyrimidine (226 mg, 1.35 mmol) andbenzyl (1s,4s)-4-aminocyclohexylcarbamate (335 mg, 1.35 mmol) in CH₃CN(6 mL), DIEA (0.600 mL, 3.45 mmol) was added. The mixture was stirred atroom temperature for 20 h. Water and EtOAc were added. The organic phasewas separated, washed with 1N HCl, dried over Na₂SO₄, concentrated invacuo to give benzyl(1s,4s)-4-(2-chloro-5-fluoropyrimidin-4-ylamino)cyclohexylcarbamate (511mg).

A solution of benzyl(1s,4s)-4-(2-chloro-5-fluoropyrimidin-4-ylamino)cyclohexylcarbamate (170mg, 0.450 mmol) and 6-amino-3,4-dihydroquinolin-2(1H)-one (109 mg, 0.670mmol) in nBuOH (3 mL) was stirred at 116° C. for 20 h. It wasconcentrated in vacuo. The residue was dissolved in MeOH (4 mL). Pd—C(10%, 30 mg) was added. The mixture was then hydrogenated under balloonhydrogen for 20 h. It was filtered through celite, and the filtrate wasconcentrated in vacuo. The residue was purified by HPLC to give thetitled compound (30 mg). MS 371.2 (M+H); UV 215.8, 263.8 nm

Example 1414-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)benzamide

The titled compound was synthesized analogously as compound6-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one,by using 4-aminobenzamide. MS 345.3 (M+H); UV 212.8, 277.8 nm

Example 1421-(4-(4-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)phenyl)piperazin-1-yl)ethanone

The titled compound was synthesized analogously as compound6-(4-((1s,4s)-4-aminocyclohexylamino)-5-fluoropyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one,by using 1-(4-(4-aminophenyl)piperazin-1-yl)ethanone. MS 428.3 (M+H); UV215.8, 252.8 nm

Example 1436-(4-((1s,4s)-4-aminocyclohexylamino)-5-(pyridin-4-yl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

To a mixture of 2,4-dichloro-5-bromopyrimidine (244 mg, 1.07 mmol) andbenzyl (1s,4s)-4-aminocyclohexylcarbamate (266 mg, 1.07 mmol) in CH₃CN(5 mL), DIEA (0.373 mL, 2.14 mmol) was added. The mixture was stirred atroom temperature for 20 h. Water and EtOAc were added. The organic phasewas separated, washed with 1N HCl, then with 5% NaHCO3, dried overNa2SO4, concentrated in vacuo to give benzyl(1s,4s)-4-(5-bromo-2-chloropyrimidin-4-ylamino)cyclohexylcarbamate (440mg).

To a mixture of benzyl(1s,4s)-4-(5-bromo-2-chloropyrimidin-4-ylamino)cyclohexylcarbamate (220mg, 0.500 mmol), pyridine-4-boronic acid (68 mg, 0.552 mmol) andPd(Ph₃P)₂Cl₂ (35 mg, 0.050 mmol) in dioxane (3 mL), aq. Na₂CO₃ (160 mg,1.50 mmol) (1.0 mL) was added. The mixture was stirred at 100° C. for 20h. More pyridine-4-boronic acid (68 mg, 0.552 mmol) and Pd(Ph₃P)₂Cl₂ (35mg, 0.050 mmol) were added. It was stirred at 100° C. for another 90min. It was then concentrated in vacuo. The residue was purified by HPLCto give benzyl(1s,4s)-4-(2-chloro-5-(pyridin-4-yl)pyrimidin-4-ylamino)cyclohexylcarbamate(93 mg).

A solution of benzyl(1s,4s)-4-(2-chloro-5-(pyridin-4-yl)pyrimidin-4-ylamino)cyclohexylcarbamate(93 mg, 0.21 mmol) and 6-amino-3,4-dihydroquinolin-2(1H)-one (52 mg,0.32 mmol) in nBuOH (4 mL) was stirred at 116° C. for 20 h. It wasconcentrated in vacuo. The residue was dissolved in MeOH (6 mL). Pd—C(10%, 44 mg) was added. The mixture was then hydrogenated under balloonhydrogen for 2 h. More Pd—C (10%, 26 mg) was added. It was stirred underballoon hydrogen for another 20 h. It was filtered through celite, andthe filtrate was concentrated in vacuo. The residue was purified by HPLCto give the titled compound (70 mg). MS 430.4 (M+H); UV 210.1, 259.6,324.8 nm

Example 1446-(4-((1s,4s)-4-aminocyclohexylamino)-5-cyclopropylpyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one

A mixture of benzyl(1s,4s)-4-(5-bromo-2-chloropyrimidin-4-ylamino)cyclohexylcarbamate (220mg, 0.500 mmol), cyclopropylboronic acid (52 mg, 0.605 mmol), K3PO4 (310mg, 1.46 mmol) and tricyclohexyl phosphine (25 mg, 0.089 mmol) intoluene (3 mL) and H2O (0.2 mL) was degassed with argon, then Pd(OAc)₂(10 mg, 0.044 mmol) was added. The mixture was stirred at 100° C. for 20h. Water and EtOAc were added. The organic phase was separated, washedwith brine, dried over Na2SO4, concentrated in vacuo to give benzyl(1s,4s)-4-(2-chloro-5-cyclopropylpyrimidin-4-ylamino)cyclohexylcarbamate(200 mg).

A solution of benzyl(1s,4s)-4-(2-chloro-5-cyclopropylpyrimidin-4-ylamino)cyclohexylcarbamate(200 mg, 0.500 mmol) and 6-amino-3,4-dihydroquinolin-2(1H)-one (105 mg,0.648 mmol) in nBuOH (4 mL) was stirred at 116° C. for 20 h. It wasconcentrated in vacuo. The residue was purified by HPLC to give benzyl(1s,4s)-4-(5-cyclopropyl-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-ylamino)cyclohexylcarbamate(97 mg).

A mixture of benzyl(1s,4s)-4-(5-cyclopropyl-2-(2-oxo-1,2,3,4-tetrahydroquinolin-6-ylamino)pyrimidin-4-ylamino)cyclohexylcarbamate(70 mg, 0.13 mmol) and Pd—C (10%, 30 mg) in MeOH (5 mL) was hydrogenatedunder balloon H2 for 20 h. It was filtered through celite. The filtratewas concentrated in vacuo. The residue was purified by HPLC to give thetitled compound (40 mg). MS 393.4 (M+H); UV 219.3, 260.8 nm

Example 145

This example illustrates methods for evaluating the compounds of theinvention, along with results obtained for such assays. The in vitro andin vivo human syk activities of the inventive compounds can bedetermined by various procedures known in the art, such as a test fortheir ability to inhibit the activity of human plasma syk. The potentaffinities for human syk inhibition exhibited by the inventive compoundscan be measured by an IC₅₀ value (in nM). The IC₅₀ value is theconcentration (in nM) of the compound required to provide 50% inhibitionof human syk proteolytic activity. The smaller the IC₅₀ value, the moreactive (potent) is a compound for inhibiting syk activity.

An in vitro assay for detecting and measuring inhibition activityagainst syk is as follows:

Inhibition of Syk Tyrosine Phosphorylation Activity

Potency of candidate molecules for inhibiting syk tyrosinephosphorylation activity is assessed by measuring the ability of a testcompound to inhibit syk-mediated tyrosine phosphorylation of asyk-specific substrate.

SYK tyrosine phosphorylation activity is measured using the LANCE™Technology developed by Perkin Elmer Life and Analytical Sciences(Boston, Mass.). LANCE™ refers to homogeneous time resolved fluorometryapplications using techniques such as time-resolved fluorescenceresonance energy transfer assay (TR-FRET) (see generally for proceduresin Perkin Elmer Application Note—How to Optimize a Tyrosine Kinase AssayUsing Time Resolved Fluorescence-Based LANCE Detection,wwww.perkinelmer.com/lifesciences). The assay principle involvesdetection of a phosphorylated substrate using energy transfer from aphosphospecific europium-labeled antibody tostreptavidin-allophycocyanin as an acceptor.

To test the ability of candidate molecules to inhibit SYK tyrosinephosphorylation activity, molecules are reconstituted in 30% DMSO andserially diluted 1:3 with the final dilution containing DMSO in theabsence of the candidate molecule. The final DMSO concentration in theassay is 3%. Kinase assays are performed as a two part reaction. Thefirst reaction is a kinase reaction and which comprises of a candidatemolecule, full length active recombinant SYK enzyme (Millipore, Calif.)and biotin-labeled SYK-specific substrate biotin-DEEDYESP-OH. The secondreaction involves termination of the kinase reaction and thesimultaneous addition of the detection reagents—europium-labeledanti-phosphotyrosine reagent (Eu-W1024-PY100, Perkin Elmer, Boston,Mass.) and Streptavidin-Allophycocyanin detection reagent (SA-APC,Prozyme, Calif.). The kinase reaction is performed in a black U-bottom96-well microtitre plate. The final reaction volume is 50 μL andcontains a final concentration of 1 nM active SYK enzyme, 550 nMSYK-substrate, and 100 μM ATP diluted in a buffer containing 50 mM TrispH 7.5, 5 mM MgCl₂, and 1 mM DTT. The reaction is allowed to proceed for1 hour at room temperature. The quench buffer contains 100 mM Tris pH7.5, 300 mM NaCl₂, 20 mM EDTA, 0.02% Brij35, and 0.5% BSA. The detectionreagents are added to the reaction mixture at the followingdilutions-1:500 for Eu-W1024-PY100 and 1:250 for SA-APC. The kinasereaction is terminated by the addition of 50 μL quench buffer containingthe detection reagents. The detection is allowed to proceed for 1 hr atroom temperature. Detection of the phosphorlated substrate in theabsence and presence of inhibitors is measured in the TR-FRETinstrument, Analyst HT (Molecular Probes, Sunnyvale, Calif.) and thecondition for measurements are set up using CriterionHost Release 2.0(Molecular Probes, Sunnyvale, Calif.). The settings used are a follows:excitation 360 nm, emission 665-7.5 nm, beam splitter 350 nm 50/50,flash 100 pulses, delay 60 us, integration 400 us, z-height 2 mmInhibition of SYK-tyrosine kinase activity is calculated as the maximumresponse observed in the presence of inhibitor, compared to that in theabsence of inhibitor. IC₅₀s were derived by non-linear regressionanalysis.

Intracellular phospho-flow cytometry was used to test compoundinhibition of Syk activity in intact non-Hodgkin's lymphoma cell linesRamos and SUDHL-6. 10×10⁶ cells in log phase growth were aliqoted; Sykkinase is activated by incubating cells for 10 minutes with 3 μg/mlantibody specific to the B cell receptor. Directly following, cells arefixed in 1% paraformaldehyde for 5 minutes at room temperature, washedin phosphate buffered saline, and then permeablized by incubation for 2hours in ice cold methanol. Cells are again washed in phosphate bufferedsaline, then incubated for 30 minutes with antibody specific forphosphorylated Erk (Y204) and BLNK (Y84), which are indicators of Sykkinase activity, and phosphorylated Syk (Y352), a measure of Src familykinase activity. All antibodies used are purchased from BD Pharmingen(San Jose, Calif.). After incubation with antibodies, cells are againwashed and subjected to flow cytometry.

The anti-proliferative effects of compounds on non-Hodgkin's lymphoma Bcell lines SUDHL-4, SUDHL-6, and Toledo was also assessed. SUDHL-4 andSUDHL-6 require B cell receptor signaling for growth and survival, whilethe Toledo cell line (serving here as a negative control) does not.Cells were aliquoted into each well of a 96-well plate and incubatedwith increasing concentrations of compound for 72 hours, after whichcell survival and proliferation was determined using the MTT assay(Chemicon International, Inc., Temecula, Calif.) following protocolssupplied by the manufacturer.

Induction of apoptosis in non-Hodgkin's lymphoma B cell lines SUDHL-4,SUDHL-6, and Toledo was assessed by measuring the apoptotis markerCaspase 3. Cells were incubated with 1, 3, or 10 μM compound for 24, 48,and 72 hours. At the conclusion of each time point, cells were processedfor flow cytometry analysis using the Monoclonal Rabbit Anti-ActiveCaspase-3 Antibody Kit and related protocols (BD Pharmingen). Data fromtwo independent experiments are presented in Tables 5A and 5B,representing the percent of total cells undergoing apoptosis followingincubation with compounds under the indicated conditions.

Syk activity is not only required for B cell signaling, proliferation,and survival, as shown, but is also critical for cellular activationupon cross-linking of the B cell receptor. B cell activation leads toincreased cell surface expression of several proteins involved in cellsignaling, antigen presentation, and adhesion. Among these, CD80, CD86,and CD69 are commonly measured to determine B cell activation status.Therefore, primary mouse B cells isolated from spleen were aliquoted andincubated with increasing concentrations of compound (0.05 to 2 μM) inthe presence of goat anti-mouse IgD (eBiosciences, Inc., San Diego,Calif.) for 20 hours to cross-link the B cell receptor. Following, cellswere washed and incubated for 30 minutes on ice with antibodies specificfor the CD80, CD86, and CD69 B cell activation markers. B cells wereidentified from the pooled population by staining with the B cell markerCD45RO. All antibodies were purchased from BD Pharmingen.

In the table below, activity in the Syk and/or Jak assays is provided asfollows: +++++=IC₅₀<0.0010 μM; ++++=0.0010 μM<IC₅₀<0.010 μM, +++=0.010μM<IC₅₀<0.10 μM, ++=0.10 μM<IC₅₀<1 μM, +=IC₅₀>1 μM.

TABLE 3 Example No. MW MH+ Syk IC50 code 1    356.393 357.1, 358.1 +++ 2   442.527 443.34, 444.47 ++ 3    356.393 357.2, 358.4 +++ 4    468.565469.36, 470.31 +++ 5 + 6 + 7 ++ 28    472.565 473.4 + 37    413.457414.3 ++ 38    428.512 429.3 + 39    371.416 372.3 ++ 40    330.355331.1 +++ 41    405.506 406.2 ++ 42    347.382 348.1 ++ 43    382.431383.2 ++ 44    525.661 526.47 + 45    473.585 475 ++ 46    473.585 474.5+++ 47    413.485 414.3 +++ 48    411.469 412.3 +++ 49    398.273 398.3;400.3; Br pattern ++++ 50    353.817 354.0, 356.0 ++++ 51    335.371336.1 ++ 52    442.542 443.1 +++ 53    486.595 487 ++ 54    415.476416.1 +++ 55    430.487 431 +++ 56    378.48 379.2 +++ 57    448.531449 + 58    348.414 349 ++ 59    333.399 334.4 + 60    350.43 351.4 + 61   392.467 393.5 + 62    367.457 368.2 +++ 63    393.495 394.2 + 64   339.403 340.2 ++ 65    381.484 382.2 ++ 66    448.575 449.3 +++ 67   460.385 460.1, 462.1 + 68    451.509 452 ++ 69    434.5 435.2, 436.2+++ 70    406.446 407.0, 408.0 +++ 71    476.585 477.0, 478.0 ++ 72   366.425 367.0, 368.0 ++ 73    352.398 352.9, 354.1 ++ 74    366.425367.0, 368.0 +++ 75    352.398 352.8, 353.8 +++ 76    371.463 372.1,373.1 ++ 77    371.463 372.2, 373.1 ++ 78    428.456 429.1, 430.2 +++ 79   265.28 266.1, 267.1 ++ 80    376.464 377.2, 378.2 ++ 81    362.437363.1, 364.1 ++ 82    336.399 337.1, 338.1 ++ 83    322.332 323.2, 324.2++ 84    393.495 394.3, 395.3 ++ 85    407.522 408.4, 409.4 ++ 86   380.452 381.2, 382.2 +++ 87    366.425 367.2, 368.2 +++ 88    331.343332.1, 333.1 ++ 89    323.36 324.2, 325.2 ++ 90    409.494 410.2, 411.2++ 91    309.333 310.2, 311.1 +++ 92    352.402 353.21 +++ 93    438.536439.07, 440.49 ++ 94    424.509 425.07, 426.32 ++ 95B    428.5 429.1 ++95A    470.537 471.1 ++ 96B    464.552 465.1 +++ 96A    506.589 507.1 ++97    401.474 402.3 ++ 98B    367.457 368.5 ++ 98A    409.494 410.5 ++99B    367.457 368.5 ++ 99A    409.494 410.5 ++ 100B    427.531 428.5 +100A    469.468 470.5 + 101    385.431 386.2 +++ 102    346.35 347.2 +103    359.393 360.3 ++++ 104    333.355 334.3 +++ 105    361.409 362.3+++ 106    369.407 370.3 ++++ 107    416.489 417.4 +++ 108    377.408378.4 + 109    418.461 419.3 + 110    418.461 419.3 + 111    425.452426.4 ++ 112    479.519 480.5 ++ 113    402.433 403.4 + 114    475.556476.4 ++ 115    423.48 424.3 ++ 15    449.56 450.6 +++ 116    520.64521.4 + 117B    422.49 423.3 + 117A    464.53 465.3 + 118    463.55464.3 ++ 119    476.581 477.3 + 120    420.473 421.3 + 121    419.489420.3 ++ 122    443.983 444.3 +++ 123    413.457 414.3 + 124    399.474400.3 + 125 442,499 443.3 + 126 455,538 456.3 ++ 127 547,635 548.4 + 128413,501 414.3 + 129 456,526 457.3 + 130 495,588 496.5 + 131 345,378346.2 +++ 132 431,343 431.4, 433.3 +++ 133 372,448 373.3 + 134 456,522457.4 ++ 135 428,468 429.3 + 136 480,548 481.5 + 137 466,521 467.5 + 138429,528 430.4 + 139 392,507 393.5 ++ 140 370,432 371.2 ++ 141 344,394345.3 ++ 142 427,528 428.3 ++ 143 429,528 430.4 + 144 392,507 393.4 +++

Inhibition of Gpvi-Mediated Platelet Function In Vitro

The ability for candidate molecules to inhibit syk-mediated plateletfunctions are tested by measuring the inhibition the GPVI-specificagonist Convulxin-induced human platelet calcium-mobilization oraggregation. Calcium-mobilization is assessed in human washed plateletsin a 96-well microtiter format. Aggregation is assessed in a 96-wellmicrotiter assay (see generally the procedures in Jantzen, H. M. et al.(1999) Thromb. Hemost. 81:111-117) or standard cuvette lighttransmittance aggregometry using human platelet-rich plasma (PRP).

Inhibition of Convulxin-Mediated Platelet Calcium-Mobilization In Vitro

Inhibition of Convulxin-induced calcium-mobilization was determined inhuman washed platelets using the FLIRP Calcium 3 Assay Kit (MolecularDevices, Sunnyvale, Calif.). For preparation of washed platelets, humanvenous blood is collected from healthy, drug-free volunteers into ACD(85 mM sodium citrate, 111 mM glucose, 71.4 mM citric acid) containingPGI₂ (1.25 ml ACD containing 0.2 μM PGI₂ final; PGI₂ was from Sigma, St.Louis, Mo.). Platelet-rich plasma (PRP) is prepared by centrifugation at160×g for 20 minutes at room temperature. Washed platelets are preparedby centrifuging PRP for 10 minutes at 730 g and resuspending theplatelet pellet in CGS (13 mM sodium citrate, 30 mM glucose, 120 mMNaCl; 2 ml CGS/10 ml original blood volume). After incubation at 37° C.for 15 minutes, the platelets are collected by centrifugation at 730 gfor 10 minutes and resuspended at a concentration of 3×10⁸ platelets/mlin Hepes-Tyrode's buffer (10 mM Hepes, 138 mM NaCl, 5.5 mM glucose, 2.9mM KCl, 12 mM NaHCO₃, pH 7.4). This platelet suspension is kept >45minutes at room temperature before use in calcium mobilization assays.

For 96-well plate Calcium-mobilization experiments, equal volumes of3×10⁸ washed platelets/ml were incubated with equal volumes of Calcium-3Assay Reagent A resuspended in 1× Hank's Balanced Salt Solution, pH 7.4,20 mM Hepes buffer. The total reaction volume of 0.2 ml/well includes1.5×10⁸/ml washed platelet/Calcium-3 Assay reagent A mix, 10 μMEptifibatide (Millennium Pharmaceuticals Inc, Cambridge, Mass.), serialdilutions (1:3) of test compounds in 0.75% DMSO. DMSO alone is added to1 well of each 8 set to allow for a maximal calcium-mobilizationreading. After 20 minutes preincubation at room temperature the 96-wellmicroplate reader is loaded into the FlexStation (Molecular Devices,Sunnyvale, Calif.). The FlexStation experimental conditions formeasuring Calcium mobilization are set up using SOFTMax Pro. Thesettings used are detailed below. Fluorescence parameters-assay mode:flex, excitation 485 nM, 525 nM with a cut-off of 515 nM; Parameters-PMTsensitivity-6, pipette height 230 μl, read time 2 minutes and 40seconds, read intervals 2 seconds, temperature-23-25° C. After 18seconds of baseline reading, calcium-mobilization is initiated by theaddition of Convulxin to a final concentration of 125 ng/ml. Inhibitionof calcium-mobilization was calculated as the maximum response observedin the presence of inhibitor, compared to that in the absence ofinhibitor. IC₅₀s were derived by non-linear regression analysis.

Inhibition of Convulxin-Mediated Platelet Aggregation In Vitro

For preparation of human platelet-rich plasma for aggregation assays,human venous blood was collected from healthy, drug-free volunteers into0.38% sodium citrate (0.013 M, pH 7.0 final). Platelet-rich plasma (PRP)is prepared by centrifugation of whole blood at 160×g for 20 minutes atroom temperature. The PRP layer is removed, transferred to a new tube,and the platelet count is adjusted, if advantageous, to achieve aplatelet concentration of ˜3×10⁸ platelets/ml using platelet-poor plasma(PPP). PPP is prepared by centrifugation of the remaining blood sample(after removal of PRP) for 20 minutes at 800×g. This preparation of PRPcan subsequently be used for aggregation assays in either a 96-wellplate or standard cuvette aggregometry.

Inhibition of Convulxin-induced aggregation is determined in 96-wellflat-bottom microtiter plates using a microtiter plate shaker and platereader similar to the procedure described by Frantantoni et al., Am. J.Clin. Pathol. 94, 613 (1990). All steps are performed at roomtemperature. For 96-well plate aggregation using platelet-rich plasma(PRP), the total reaction volume of 0.2 ml/well includes 190 μl of PRP(˜3×10⁸ platelets/ml, see above), and 5 μl of either serial dilution oftest compounds in 30% DMSO or buffer (for control wells). After 20minutes preincubation at room temperature 5 μl of 320 ng/ml Convulxinagonist solution is added to each well to give a final concentration of8 ng/ml Convulxin. The plates are then agitated for 5 min on amicrotiter plate shaker and the 5 minute reading is obtained in themicrotitre plate reader (Softmax, Molecular Devices, Menlo Park,Calif.). Aggregation is calculated from the decrease of OD at 650 nm att=5 minutes. IC₅₀s were derived by non-linear regression analysis.

Inhibition of Convulxin-induced aggregation was also determined forcuvette light transmittance aggregation assays, serial dilutions (1:2)of test compounds were prepared in 30% DMSO in a 96 well V-bottom plate(final DMSO concentration in the cuvette was 0.3%). The test compound (5μl of serial dilutions in DMSO) was preincubated with PRP for 20 minutesprior to initiation of aggregation reactions, which is performed in aChronoLog aggregometer by addition of agonist (125-250 ng/ml Convulxin)to 495 μL of PRP at 37° C. The aggregation reaction is recorded for 4min, and maximum extent of aggregation is determined by the differencein extent of aggregation at baseline, compared to the maximumaggregation that occurs during the 4 minute period of the assayInhibition of aggregation was calculated as the maximum aggregationobserved in the presence of inhibitor, compared to that in the absenceof inhibitor. IC₅₀s were derived by non-linear regression analysis.

The following table 4 gives syk and PRP IC₅₀ values.

Calcium Flux Assay in Ramos Cells Induced by BCR Cross-Linking

Ramos cells (2G6.4C10, Burkitt's lymphoma, ATCC Item Number: CRL-1923)are sub-cultured at 5×10⁵ cells/ml in fresh medium 3 or 4 days ahead ofexperiments. Cells are harvest and re-suspend in fresh medium at 8×10⁶cells/ml before dye-loading. An equal volume of Calcium 3 loading dye(Molecular Device) is added and mixed into cell suspension. Loadingcells are dispensed in a 96 well plate and incubated 30 min. Compoundsare then added in the dye-loaded cells and incubated for another 30 min.Spin cell down at 1000 rpm for 3 min before fluorescence measurement inFlexStation. BCR stimulation is carried by the addition of 5 μg/mlantibody (AffiniPure F(ab′)₂ fragment Donkey anti-human IgM, JacksonImmunoResearch Laboraotries).

Calcium Flux Assay in Jurkat Cells Induced by TCR Cross-Linking

The protocol is very similar to B cell calcium flux as described in theprevious section. The only differences are that T cells (clone E6-1,Acute T cell Leukemia, ATCC Item Number: Tib-152) and anti-human CD3(Functional Grade Purified anti-human CD3, clone OKT3, eBioscience, No.16-0037) replaced B cells and anti-human IgM. Cell density is kept thesame but antibody is used at a concentration of 100 ng/ml.

IL-2 Secretion in Jurkat Cells Induced by TCR Cross-Linking

Jurkat cell propagation and compound incubation procedures are the sameas described in Jurkat calcium flux assay in the previous section.Antibody (anti CD3, OKT3) is coated onto a fresh plate (without cells)at 100 ng/well. Cells are suspended at 8×10⁶ cells/ml and incubated withcompounds for 30 min in a separate plate. At the end of incubation,cells are transferred to the antibody-coated plate and incubated for 16hours. 100 μl of cell medium after incubation is used for IL-2measurement after incubation. IL-2 level is determined using an IL-2ELISA kit (Human IL-2 ELISA kit II, BD Bioscience, No. 550611).

TABLE 4

Ex syk PRP No Structure MW MS IC50 IC50 Name  8

420.521 421.5 ++ 25.2675 1-(2-(4-(piperazin- 1-yl)phenylamino)-7H-pyrrolo[2,3-d] pyrimidin-4- yl)piperidine-3- carboxamide  9

462.558 463.6 ++ 12.297 1-(2-(4-(4- acetylpiperazin-1- yl)phenylamino)-7H-pyrrolo[2,3- d]pyrimidin-4- yl)piperidine-3- carboxamide 10

406.538 407.5 ++++ 33.167 4-(4-(aminomethyl) piperidin-1-yl)-N-(4-(piperazin-1- yl)phenyl)-7H- pyrrolo[2,3-d] pyrimidin-2-amine 11

448.575 449.5 ++++  5.281 1-(4-(4-(4-(4- (aminomethyl)piperidin-1-yl)-7H- pyrrolo[2,3-d] pyrimidin-2- ylamino)phenyl)piperazin-1- yl)ethanone

348.414 349.4, 350.4 + (S)-4-(2-(amino- methyl)pyrrolidin- 1-yl)-N-(1H-indazol-6-yl)-7H- pyrrolo[2,3-d] pyrimidin-2- amine 12

379.468 380.5 + (S)-1-(2-(4- (piperazin-1-yl) phenylamino)-7H-pyrrolo[2,3-d] pyrimidin-4- yl)pyrrolidin-3-ol 13

421.505 422.5 + (S)-1-(4-(4-(4-(3- hydroxypyrrolidin- 1-yl)-7H-pyrrolo[2,3-d]pyrimidin- 2-ylamino) phenyl)piperazin- 1-yl)ethanone 14

473.585 474.5 +++ 50 2-(4-(4-acetyl- piperazin-1-yl) phenylamino)-4-(4-(aminomethyl) piperidin-1-yl)-7H- pyrrolo[2,3-d] pyrimidine-5-carbonitrile

453.547 454.6 ++ 2-(4-(4-acetyl- piperazin-1- yl)phenylamino)-4-(3-(hydroxy- methyl)piperidin-1- yl)pyrimidine-5- carboxamide

467.574 468.6 + 2-(4-(4-acetyl- piperazin-1-yl) phenylamino)-4-(2-(2-hydroxyethyl) piperidin-1- yl)pyrimidine-5- carboxamide 15

449.563 450.6 +++ 50 1-(4-(4-(6-(4- (aminomethyl) piperidin-1-yl)-9H-purin-2-ylamino) phenyl)piperazin-1- yl)ethanone

348.414 349.4, 350.4 ++ (S)-4-(3-amino- piperidin-1-yl)-N-(1H-indazol-6- yl)-7H-pyrrolo[2,3- d]pyrimidin-2- amine

434.548 435.41, 436.54 ++ 13.75 (S)-1-(4-(4-(4-(3- aminopiperidin-1-yl)-7H-pyrrolo[2,3- d]pyrimidin-2- ylamino)phenyl) piperazin-1-yl)ethanone 16

341.394 342.5 +++  0.2955 N-(4-(4-(amino- methyl)piperidin-1-yl)-5-fluoro- pyrimidin-2-yl)- 1H-indazol-6- amine 17

391.447 392.5 ++ 4-(4-(aminomethyl) piperidin-1-yl)-5- fluoro-N-(3,4,5-trimethoxy- phenyl)pyrimidin- 2-amine 18

427.528 428.6 +++  0.195 1-(4-(4-(4-(4- (aminomethyl) piperidin-1-yl)-5-fluoropyrimidin-2- ylamino)phenyl) piperazin-1- yl)ethanone 19

377.496 378.5 + N-(4-(piperazin-1- yl)phenyl)-4- (piperidin-1-yl)-7H-pyrrolo[2,3-d] pyrimidin-2-amine 20

419.533 420.5 + 1-(4-(4-(4- (piperidin-1-yl)- 7H-pyrrolo[2,3-d]pyrimidin-2- ylamino)phenyl) piperazin-1-yl) ethanone 21

363.469 364.5 ++ N-(4-(piperazin-1- yl)phenyl)-4- (pyrrolidin-1-yl)-7H-pyrrolo[2,3- d]pyrimidin-2- amine 22

405.506 406.5 ++ 1-(4-(4-(4- (pyrrolidin-1-yl)- 7H-pyrrolo[2,3-d]pyrimidin-2- ylamino)phenyl) piperazin-1-yl) ethanone 23

420.521 421.5 + 1-(2-(4-(piperazin- 1-yl)phenylamino)- 7H-pyrrolo[2,3-d]pyrimidin-4- yl)piperidine-4- carboxamide 24

462.58 463.5 ++ 1-(2-(4-(4- acetylpiperazin-1- yl)phenylamino)-7H-pyrrolo[2,3- d]pyrimidin-4- yl)piperidine-4- carboxamide 25

452.563 453.6 + 2-(4-(4-acetyl- piperazin-1-yl) phenylamino)-4-(4-(aminomethyl) piperidin-1- yl)pyrimidine-5- carboxamide 26

355.377 356.3 ++ 1-(2-(1H- indazol-6- ylamino)-5- fluoropyrimidin-4-yl)piperidine-3- carboxamide 27

441.511 442.4 + 1-(2-(4-(4- acetylpiperazin-1- yl)phenylamino)-5-fluoropyrimidin- 4-yl)piperidine-3- carboxamide

482.589 483.6 ++ 2-(4-(4-acetyl- piperazin-1-yl) phenylamino)-4-(4-(2-hydroxyethyl)- 1,4-diazepan-1- yl)pyrimidine-5- carboxamide

472.565 473.4 + tert-butyl (1-(5-fluoro-2-(4- (N-methyl-acetamido)phenyl- amino)pyrimidin- 4-yl)piperidin-4- yl)methylcarbamate30

444.511 445.4 + tert-butyl (1-(2-(4-carbamoyl- phenylamino)-5-fluoropyrimidin-4- yl)piperidin-4- yl)methylcarbamate 31

344.394 345.3 +++  0.3455 4-(4-(4-(amino- methyl)piperidin-1-yl)-5-fluoro- pyrimidin-2- ylamino)benzamide 32

380.446 381.3 +++  4.6705 4-(4-(4-(amino- methyl)piperidin-1-yl)-5-fluoro- pyrimidin-2- ylamino)benzene- sulfonamide 29

372.448 373.3 +++  0.3155 N-(4-(4-(4- (aminomethyl) piperidin-1-yl)-5-fluoropyrimidin-2- ylamino)phenyl)- N-methylacetamide 33

370.432 371.3 +++  0.3355 6-(4-(4-(amino- methyl)piperidin-1-yl)-5-fluoro- pyrimidin-2- ylamino)-3,4- dihydroquinolin- 2(1H)-one 34

427.53 428.4 + 1-(4-(4-(4- (2-(aminomethyl) piperidin-1-yl)-5-fluoropyrimidin-2- ylamino)phenyl) piperazin-1-yl) ethanone 35

441.51 442.3 + tert-butyl (1-(2-(1H- indazol-6- ylamino)-5-fluoropyrimidin-4- yl)piperidin-2- yl)methylcarbamate 36

341.39 342.3 + N-(4-(2-(amino- methyl)piperidin- 1-yl)-5-fluoro-pyrimidin-2-yl)-1H- indazol-6-amine

Example 146 Millipore Upstate KinaseProfiler™ Screening

This assay is a direct measurement of the effect of compound on thecatalytic activity of JAK3. Purified human JAK3 (GenBank AF513860)sequence (residue 781-C terminus) was obtained from insect cells. Thecatalytic hydrolysis of ATP is measured using a radiometric filterbinding method. Incubation of kinase with ³³-[P]ATP and substrate leadsto incorporation of ³³[P] into the substrate which can then be separatedfrom the other reaction components by filtration. Assays were performedusing 10 μM ATP and in the absence or presence of 1, 0.3, or 0.1 μMcompound. Activity was expressed as % of inhibition of control.

In the table below, activity in the Jak assays is provided as follows:+++++=IC₅₀<0.0010 μM; ++++=0.0010 μM<IC₅₀<0.010 μM, +++=0.010μM<IC₅₀<0.10 μM, ++=0.10 μM<IC₅₀<1 μM, +=IC₅₀>1 μM.

TABLE 5 Inhibition (IC50%) of catalytic activity of JAK1, 2 and 3 by thecompound Concentration (μM) Compound JAK 1 JAK 2 JAK 3

++ ++ ++

+ ++ ++

+ ++ ++

+ + +

++ ++ ++

+++ +++ +++

+++ +++ +++

+ ++ +

+++ +++ +++ Inhibition (%) of catalytic activity of JAK3 by 1, 0.3 or0.1 μM compound nd Concentration (μM) 1 μM 0.3 μM 0.1 μM

77 41 ND

ND 82 ND ND: not done

Example 147 Ambit KinomeScan Screening

This assay is an ATP-site dependent competition binding assay in whichhuman kinases of interest are fused to a proprietary tag (T7bacteriophage). The amount of kinase bound to an immobilized,active-site directed ligand is measured in the presence and absence ofthe test compound. Ambit's JAK assays use kinase domains and notfull-length proteins. The domain used for JAK1 binding is the pseudokinase domain while that for JAK3 binding is the catalytic domain (MazenW Karaman, Sanna Herrgard, Daniel K Treiber, et. al. A Quantitativeanalysis of kinase inhibitotr selectivity. Nature Biotechnology, 2008,Volume 26, No. 1, Page 127-132).

Example 148 JAK3/STAT6 Cellular Assay

Stimulation of Ramos B cells by interleukin 4 (IL4) leads to signalingthrough JAK1/JAK3 resulting in phosphorylation of STAT6 (signaltransducers and activators of transcription). The effect of compounds oninhibition of JAK3 and/or JAK1 can be assessed by measuring the amountof phosphorylated STAT6. This is performed by Western blotting using aspecific phospho-STAT6 antibody.

Ramos B cells were suspended in 10 mM Hepes-buffered RPMI media (2×10⁷cells/ml). Cells (90 μl) were incubated with 10 μl 3.3 μg/ml interleukin4 (R & D Systems Inc, cat #204-IL; final concentration: 0.33 μg/ml).Incubations were for 10 min at 37° C. in the absence or presence of 2 μlcompound diluted in 30% DMSO. Reactions were terminated by the additionof an equal volume of 2× lysis buffer (100 mM TRIS-HCl pH 8.0, 2%Triton-X-100, 5 mM EDTA, 250 mM NaCl, 20% glycerol, 1.25 mM PMSF, 5 mMsodium orthovandate, 5 mM (β-glycerophosphate, mini complete EDTAprotease inhibitor cocktail (Sigma)).

Samples were incubated with 1 μl of the nuclease, benzonase (Novagen,cat #71205-3) for 1 hour, room temperature and then 50 μl 5× loadingbuffer (330 mM TRIS pH 6.8, 9.5% SDS, 34% glycerol, 0.01% bromophenolblue, 10% beta-mercaptoethanol) was added.

Cell lysates (15 μL) were subjected to SDS-PAGE (Novex 4-12%TRIS-glycine gels, Invitrogen) under reducing conditions, followed byelectroblot-transfer onto nitrocellulose membranes. Membranes were thenincubated in Zymed blocking buffer (Invitrogen) for 1 hr at roomtemperature (RT) then overnight at 4° C. with 1:500 antiphosphotyrosine-STAT6 (Cell Signaling Technology, cat #9364) primaryantibody in Zymed blocking buffer. Following 5×10 min washes withTris-buffered saline, 0.25% NP40 (TBSN), blots were incubated for 1 hrat room temperature in the presence of 1:10,000 HRP-conjugated donkeyanti-rabbit secondary antibody (Amersham Biosciences, cat #NA934V) inZymed blocking buffer. After 4×10 min TBSN washes, blots were visualizedby ECL (Pierce Western Lightening, Perkin Elmer cat #NEL101). In orderto determine total β3 content, blots were stripped, washed 4× with TBSN,and re-probed with 1:2000 C3A antibody in block buffer overnight at 4°C. After 4×10 min TBSN washes, blots were incubated with 1:10,000 goatanti-mouse secondary antibody in blocking buffer, washed 4 more timeswith TBSN and exposed to Western Lightening reagent.

Levels of stimulation over background and the extent of inhibition ofcompound were determined by densitometry. FIG. 4 shows the effect ofincreasing dose of a compound of the invention on pSTAT6 formation inresponse to IL4 stimulation of B Ramos cells.

Example 149 Inhibition of Jak Kinase Activity Assay for Ramos B-CellLine Stimulated with IL-4

These examples illustrate methods for evaluating the in vitro and invivo human JAK kinase activities of the inventive compounds can bedetermined by various procedures known in the art, such as a test fortheir ability to inhibit the activity of human plasma JAK kinase. Thepotent affinities for human JAK kinase inhibition exhibited by theinventive compounds can be measured by an IC₅₀ value (in nM). The IC₅₀value is the concentration (in nM) of the compound required to provide50% inhibition of human JAK kinase activity. The smaller the IC₅₀ value,the more active (potent) is a compound for inhibiting JAK kinaseactivity.

An in vitro assay for detecting and measuring inhibition activityagainst JAK kinase is as follows:

The activity of the compounds for JAK kinases is confirmed in cellularassays designed to test for JAK inhibition. Briefly, JAK inhibition istested in human Ramos B-cells activated with cytokine Interleukin-4(IL-4). Twenty to 24 hours post stimulation, the cells are stained forupregulation of CD23 and analyzed by FACS. Stimulation of the B-cellswith IL-4 leads to the activation of the JAK/STAT pathway throughphosphorylation of the JAK kinase JAK1 and JAK3, which in turnphosphorylate and activate transcription of factors STAT-5 and STAT-6.The low-affinity IgE receptor (CD23) is upregulated by activated STAT-5.

For the assay, human Ramos B-cells (ATCC, Catalog No. CRL-1596) arecultured in RPMI 1640 medium (Cellgro, Catalog No. 10-040-CM) containing10% fetal bovine serum (JRH, Catalog No. 12106-500M) according to thepropagation protocol supplied with the cells, and maintained at adensity of approximately 3.5×10⁵ cells/ml. The day before the assay, thecells are diluted to 3.5×10⁵ cells/ml to insure they are in thelogorithmic growth phase. The cells are spun down, and suspended in RPMI1640 medium (Cellgro, MediaTech, Inc., Herndon, Va., Cat No. 10-040-CM)containing 5-10% fetal bovine serum (FBS), heat inactivated (JRHBiosciences, Inc, Lenexa, Kans., Cat No. 12106-500M) according to ATCCpropagation protocol. Cells are maintained at a density of 3.5×10⁴⁻⁵cells/ml. The day before the experiment, Ramos B-cells are diluted to3.5×10⁵ cells/mL to ensure that they are in a logarithmic growth phaseand aliquots dispensed into a 96-well tissue culture plate. Cells areincubated with test compound (dissolved in DMSO) or DMSO (control) for 1hr at 37° C. and then stimulated with IL-4 (Pepotech, Catalog No.200-04) for 20-24 hours (final concentration is 50 Units/ml).

Cells are spun down and suspended in RPMI with 5% serum. 5×10⁴ cells areused per point in a 96-well tissue culture plate. Cells arepre-incubated with compound or DMSO (Sigma-Aldrich, St. Louis, Mo., CatNo. D2650) vehicle control for 1 hour in a 37° C. incubator. Cells arethen stimulated with IL-4 (Peprotech Inc., Rocky Hill, N.J., Cat No.200-04) for a final concentration of 50 units/mL for 20-24 hours. Cellsare then spun down and stained with anti-CD23-PE(BD Pharmingen, SanDiego, Calif., Cat No. 555711) and analyzed by FACS. Detection isperformed using a BD LSR I System Flow Cytometer, purchased from BectonDickinson Biosciences of San Jose, Calif.

Proliferation is measured using CellTiter-Glo.RTM Luminescent CellViability Assay (Promega), which determines the number of viable cellsin culture based on quantitation of the ATP present, as an indicator ofmetabolically active cells. The substrate is thawed and allowed to cometo room temperature. After mixing the Cell Titer-Glo reagent and diluenttogether, 100 μL is added to each well. The plates are mixed on anorbital shaker for two minutes to induce lysis and incubated at roomtemperature for an additional ten minutes to allow the signal toequilibrate. Detection is performed using a Wallac Victor2 1420multilabel counter purchased from Perkin Elmer, Shelton, Conn.

On day two, A549 cells are pre-incubated with a 2,4-pyrimidinediaminetest compound or DMSO (control) (Sigma-Aldrich, St. Louis, Mo., CatalogNo. D2650) for 1 hour. The cells are then stimulated with IFNγ (75ng/mL) (Peprotech Inc., Rocky Hill, N.J., Cat. No. 300-02) and allowedto incubate for 24 hours. The final test compound dose range is 30 μM to14 nM in 200 μL F12K media containing 5% FBS, 0.3% DMSO.

On day three, the cell media is removed and the cells are washed with200 μL PBS (phosphate buffered saline). Each well is trypsinized todissociate the cells, then neutralized by addition of 200 μL completeF12K media. Cells are pelleted and stained with an APC conjugated mouseanti-human ICAM-1 (CD54) (BD Pharmingen, San Diego, Calif., Catalog#559771) antibody for 20 minutes at 4° C. Cells are washed with ice coldFACS buffer (PBS+2% FBS) and surface ICAM-1 expression is analyzed byflow cytometry. Detection is performed using a BD LSR I System FlowCytometer, purchased from BD Biosciences of San Jose, Calif. Events aregated for live scatter and the geometric mean is calculated(Becton-Dickinson CellQuest software version 3.3, Franklin Lakes, N.J.).Geometric means are plotted against the compound concentration togenerate a dose response curve.

Example 150 Inhibition of Syk-Mediated Signal Transduction Through the BCell Receptor in Non-Hodgkin's Lymphoma Cell Lines

Cells were pre-treated for 1 hour without or with compound (0.02 to 2uM) prior to stimulation of B cell receptor singling by incubation ofcells with 3 μg/ml anti-mu antibody for 10 minutes at 37° C. Ca²⁺ fluxwas measured using the Calcium 3 loading dye and the FlexStation(Molecular Device). B cell receptor signaling was assayed byintracellular phospho-Flow Cytometry, following protocols supplied by BDPharmingen (San Jose, Calif.). Syk activation was measured by inductionof BLNK tyrosine phosphorylation at amino acid position 84 (pBLNK Y84)and induction of ERK1/2 tyrosine phosphorylation at amino acid position204 (pERK Y204). Activation of the Src family member Lyn was measured byinduction of Syk tyrosine phosphorylation at amino acid position 352(pSyk Y352). Data are presented as μM IC₅₀s. Each compound effectivelyinhibited B cell receptor-induced Ca²⁺ fluxing and activation of Syk,but not the Src family member Lyn.

Example 151 Syk Inhibition Exerts an Anti-Proliferative Effect onNon-Hodgkin's Lymphoma Cell Lines

Cells were incubated with increasing concentrations of each compound,then evaluated at 72 hours for extent of proliferation using the MTTassay (company, city, state) following the manufacturer suppliedprotocol. Data are presented as μM IC₅₀ values, representing the meanplus/minus standard deviation from 5 or 6 independent experiments. Eachcompound inhibited proliferation of SUDHL-4 and -6 cell lines, whichrely on Syk for survival and growth signals, in the low μM range. Toledocells which do not require Syk, was noticeably less sensitive to theanti-proliferative effects of Syk inhibition.

Example 152 Syk Inhibition Induces Apoptosis in Non-Hodgkin's LymphomaCell Lines

Data represent two independent experiments to evaluate the effect of Sykand Syk/JAK inhibition on survival of diffuse large non-Hodgkin'slymphoma B cell lines. SUDHL-4 and SUDHL-6 cells lines rely onSyk-mediated B cell receptor signaling for survival, while Toledo cellsdo not. Cells are incubated with compounds at the various concentrationsand times; induction of apoptosis is measured by flow cytometry usingthe Caspase 3 Detection Kit (Sigma-Aldrich, Saint Luis, Mo.). Data ispresented as the percent of total cells positive for the apoptosismarker, caspase 3.

Example 153 Inhibition of Mouse Primary B Cell Activation by SykInhibitors

Mouse primary splenocytes were pre-treated for 1 hour with increasingconcentrations of each compound (0.05-2 μM) prior to addition of controlor goat anti-mouse IgD serum. Anti-IgD induced B cell activation wasmeasured 16 hours later by flow cytometry, staining for the activationmarkers CD80/86 and CD69. Data represent IC₅₀ ranges for the inhibitionof B cell activation.

Example 154 Mouse Model of Immune-Mediated Thrombocytopenia

Immune-mediated thrombocytopenia is caused by antibodies directedagainst platelet surface glycoproteins, antibodies againstdrug-containing complexes on the platelet surface, or by antibody-coatedcells or immune complexes that interact with the platelet surface.Select compounds were evaluated for their ability to inhibit plateletclearance in a mouse model of antibody-mediated thrombocytopenia. Inthis model, a rapid clearance of circulating platelets (approximately50%) results from the intravenous administration of a rat anti-mouseGPIIb (clone MWReg30) antibody (BD Biosciences, Pharmingen). To evaluatecapacity for inhibition of platelet clearance, compounds were suspendedinto 0.5% methycellulose in water and administered via oral gavage (100ul/mouse) at a time prior to antibody injection when the compound wouldachieve maximum plasma concentration (typically 1-2 hours based onseparate pharmacokinetic experiments for individual compounds). At 4 and8 hours after injection of antibody, terminal blood samples wereobtained from groups of vehicle and test article treated mice (n=5-10mice/group) via cardiac puncture. Blood was anticoagulated usingtrisodium citrate or EDTA. Whole blood samples were measured forplatelet counts on a hematology analyzer (Hemavet, Drew Scientific).Remaining blood was processed for plasma and compound concentrationsmeasured by mass spectrometry.

Platelet clearance was determined by measuring the difference inplatelet number between the average non-antibody treatment group andanimals administered the rat anti-mouse GPIIb antibody. Inhibition ofplatelet clearance was determined by comparing the difference betweenplatelet clearance of vehicle and compound treated animals.

Example 155 Mouse Model of Collagen Antibody Induced Arthritis

The inhibitory activity of select compounds was investigated in a mousemodel of collagen antibody induced arthritis (CAIA). Collagen inducedarthritis is mediated by autoantibodies to type II collagen andcomplement, thus arthritis can be induced by administration ofpolyclonal antibodies or a mixture of monoclonal antibodies to type IIcollagen. The CAIA model (Chondrex, Inc., Redmond, Wash.) uses a mixtureof 4 clones which recognize individual epitopes clustered within an 83amino acid peptide fragment of type II collagen. These epitopes sharecommon amino acid sequences with many different species of type IIcollagen including chicken, mouse, rat, bovine, porcine, monkey andhuman. The model utilizes a monoclonal antibody cocktail followed bybacterial lipopolysaccharide (LPS) to induce a severe and consistentarthritis in mice within 7 days. This model was developed based on thehypothesis that bacterial toxins absorbed through the gastrointestinaltract play a synergistic and pathologic role with autoantibodies to typeII collagen in triggering arthritis in patients with RheumatoidArthritis.

For these experiments, the monoclonal antibody cocktail (Lot # OC-708)was injected intravenously via tail vein at a dose of 4 mg/mouse (40mg/ml) on day 0 followed by intraperitoneal injection of LPS dilutedinto normal saline at a dose of 25 ug/mouse in 8 week old, female Balb/Cmice (Charles River, Inc.). Dosing of test articles was started justbefore or after the IV injection of antibody cocktail. Compounds weresuspended into 0.5% methylcellulose in water and administered via oralgavage (100 ul/mouse) daily for the duration of the 7-10 day study.Clinical inflammation scores were obtained daily. Inhibition of clinicalinflammation scores was determined based on the difference betweenvehicle and test article treated mice at the end of the experiment.Plasma concentrations represent peak concentration at 1 hour post lastdose on the day of study termination.

Example 156 Inhibition of IL-4 Induced Jak1/3 to Stat-6 Phosphorylationin Ramos B Cells

Ramos B cells were pre-treated for 1 hour with increasing concentrationsof compound, as indicated prior to addition of IL-4. Cells wereincubated with IL-4 for 10 minutes, and then subjected to intracellularflow cytometry to measure the percent inhibition of IL-4 induced Stat-6.

Example 157 Analysis of B Cell Signaling

The human non-Hodgkin's lymphoma B cell lines SUDHL-4 (#ACC 495),SUDHL-6 (#ACC572), and Karpas-422 (#ACC32) were obtained from DSMZ(Braunschweig, Germany); Toledo (#CRL-2631) and Ramos (#CRL-1596) wereobtained from the American

Type Culture Collection (ATCC; Manassas, Va.). All cells were maintainedin RPMI media (Invitrogen, Carlsbad, Calif.) supplemented with 10% fetalcalf serum (ATCC) and penicillin/streptomycin (Invitrogen), andmaintained in a 37° C. humidified tissue culture incubator. Antibodiesused in these studies include polyclonal goat F(ab)′2 anti-human IgG(H+L) and anti-human IgM (BioSource, Camarillo, Calif.); rabbitanti-human Syk, rabbit anti-human phospho-Syk (Y525/526), rabbitanti-human phospho-Syk (Y352), anti-human BLNK, anti-human phospho-BLNK(Y84) were obtained from Cell Signaling Technologies, Inc. (Danvers,Mass.). The following antibodies were obtained from Becton Dickenson(San Jose, Calif.) for phospho-flow cytometry: Alexa fluor488-conjugated mouse anti-human phospho-STATE (Y641), Phycoerythrin(PE)-conjugated mouse anti-human phospho-Zap70 (Y319)/Syk(Y352), andFluorescein isothiocyanate (FITC)-conjugated mouse anti-humanphospho-ERK1/2 (T202/Y204).

Phospho-flow cytometry was performed essentially as described elsewhere(Irish, Czerwinski et al. Blood 108(9): 3135-42 (2006). 0.5×10⁶ cells ingrowth media were stimulated with 1 μg/ml anti-μ or anti-γ antibody for10 minutes. Induced signaling was terminated immediately following theindicated time by the addition of paraformaldehyde (Electron MicroscopySciences, Hatfield, Pa.) to a final concentration of 1%. Cells wereincubated with paraformaldehyde for 5 minutes at room temperature,washed once with phosphate buffered saline (PBS), then resuspended andincubated overnight at 4° C. in pre-chilled methanol (−80° C.) (company,address). Fixed and permeablized cells were subsequently washed once inPBS, a second time in PBS containing 1% bovine serum albumin (BSA)(Sigma-Aldrich, St. Louis, Mo.), and then stained with the indicatedantibodies diluted 1:20 in PBS+1% BSA. After 30 minutes, cells werewashed once in PBS and subjected to flow cytometry using the FACSCalibur (Becton Dickenson). For Western blot analyses, 106 cells werestimulated for 30 minutes with 2 μg/ml of the indicated BCR-specificantibodies. Signaling was terminated by resuspending the cells in lysisbuffer and incubated on ice for 1 hour. Cell debris were removed bycentrifugation, and the cleared protein lysates were resolved by 10%SDS-PAGE and probed with the indicated antibodies followingrecommendations made by the manufacturers. Where indicated, cells werepre-treated for 1 hour at 37° C. with Syk inhibitors or vehicle control(0.5% DMSO) at several concentrations prior to stimulation with anti-BCRantibody.

Example 158 Analysis of B Cell Signaling

The human non-Hodgkin's lymphoma B cell lines SUDHL-4 (#ACC 495),SUDHL-6 (#ACC572), and Karpas-422 (#ACC32) were obtained from DSMZ(Braunschweig, Germany); Toledo (#CRL-2631) and Ramos (#CRL-1596) wereobtained from the American Type Culture Collection (ATCC; Manassas,Va.). All cells were maintained in RPMI media (Invitrogen, Carlsbad,Calif.) supplemented with 10% fetal calf serum (ATCC) andpenicillin/streptomycin (Invitrogen), and maintained in a 37° C.humidified tissue culture incubator. Antibodies used in these studiesinclude polyclonal goat F(ab)′2 anti-human IgG (H+L) and anti-human IgM(BioSource, Camarillo, Calif.); rabbit anti-human Syk, rabbit anti-humanphospho-Syk (Y525/526), rabbit anti-human phospho-Syk (Y352), anti-humanBLNK, anti-human phospho-BLNK (Y84) were obtained from Cell SignalingTechnologies, Inc. (Danvers, Mass.). The following antibodies wereobtained from Becton Dickenson (San Jose, Calif.) for phospho-flowcytometry: Alexa fluor 488-conjugated mouse anti-human phospho-STATE(Y641), Phycoerythrin (PE)-conjugated mouse anti-human phospho-Zap70(Y319)/Syk(Y352), and Fluorescein isothiocyanate (FITC)-conjugated mouseanti-human phospho-ERK1/2 (T202/Y204).

Phospho-flow cytometry was performed essentially as described elsewhere(Irish, Czerwinski et al. Blood 108(9): 3135-42 (2006). 0.5×10⁶ cells ingrowth media were stimulated with 1 μg/ml anti-μ a or anti-γ antibodyfor 10 minutes. Induced signaling was terminated immediately followingthe indicated time by the addition of paraformaldehyde (ElectronMicroscopy Sciences, Hatfield, Pa.) to a final concentration of 1%.Cells were incubated with paraformaldehyde for 5 minutes at roomtemperature, washed once with phosphate buffered saline (PBS), thenresuspended and incubated overnight at 4° C. in pre-chilled methanol(−80° C.) (company, address). Fixed and permeablized cells weresubsequently washed once in PBS, a second time in PBS containing 1%bovine serum albumin (BSA) (Sigma-Aldrich, St. Louis, Mo.), and thenstained with the indicated antibodies diluted 1:20 in PBS+1% BSA. After30 minutes, cells were washed once in PBS and subjected to flowcytometry using the FACS Calibur (Becton Dickenson). For Western blotanalyses, 106 cells were stimulated for 30 minutes with 2 μg/ml of theindicated BCR-specific antibodies. Signaling was terminated byresuspending the cells in lysis buffer and incubated on ice for 1 hour.Cell debris were removed by centrifugation, and the cleared proteinlysates were resolved by 10% SDS-PAGE and probed with the indicatedantibodies following recommendations made by the manufacturers. Whereindicated, cells were pre-treated for 1 hour at 37° C. with Sykinhibitors or vehicle control (0.5% DMSO) at several concentrationsprior to stimulation with anti-BCR antibody.

Example 159 Calcium Flux Assay and Selective Inhibition of Syk inNon-Hodgkin's Lymphoma B Cell Lines

Ramos cells were cultured (maintaining approximately 0.5×10⁶ cells/ml)in growth medium 3 to 4 days ahead of experiments. Cells were harvestedand re-suspended in fresh medium at 8×10⁶ cells/ml before dye-loading.An equal volume of Calcium 3 loading dye (Molecular Device, Sunneyvale,Calif.) was added to the cell suspensions. Loaded cells were dispensedin a 96 well plate and incubated for 20 minutes. Syk inhibitors werethen added to the loaded cells and incubated for another 30 minutes. Bcells were stimulated with 5 μg/ml anti-μ antibody. Changes inintracellular Ca2+ concentration was measured using the FlexSTATion(Molecular Devices, Sunnyvale, Calif.).

The selectivity and potency of Syk inhibition in B cells was initiallyinterrogated by Western blot, measuring BCR-mediated induction of pSykY525/526 and pBLNK Y84, both measures of Syk kinase activity, and theinduction of pSyk Y352, a measure of Src kinase activity. SUDHL-6 Bcells were stimulated with anti-BCR specific antibody for 30 minutes inthe presence or absence of each Syk inhibitor or vehicle control.Treatment with 0.16 or 1 μM of each compound reduced BCR-induced Sykautophorphorylation (Y525/526) by roughly 40% and 60%, respectively, asestimated by densitometry (data not shown). An expanded range ofconcentrations was used to further evaluate the effect of thesecompounds on BCR induced Syk and Src kinase activity.

The ability of each compound to suppress signaling events more distal tothe BCR was also measured. Cells were again stimulated by anti-BCRantibody in the presence or absence of various concentrations of eachSyk inhibitor. The induction of pSyk Y352 was measured as a specificitycontrol, while that of pERK1/2/Y204 was used as a measure of more distalSyk-dependent signaling (Jiang, Craxton et al. J Exp Med 188(7):1297-306 (1998). This experiment was repeated, in which the effect bothcompounds on Src and Syk activity were determined (FIG. 10).Concentrations of less than 125 nM were sufficient to suppress BCRinduced Syk signaling to ERK1/2. By contrast, much higher concentrationswere required to cause a modest suppression of Src activity; an effecton Src that was not observed by Western blot. None of these Sykinhibitors suppressed PMA-induced ERK1/2 tyrosine phosphorylation,demonstrating these compounds do not inhibit signaling eventsdown-stream of PKC. This shows that selective inhibition of Syksuppressed BCR-induced Ca2+ flux in B cells with IC₅₀ values around 100nM. This suggests that by inhibiting Syk, these compounds suppress thesignaling pathway, blocking the cellular response.

Example 160 Caspase 3 and Proliferation Assays: Syk Inhibition DisruptsProliferation and Survival of Non-Hodgkin's Lymphoma B Cell Lines

Induction of apoptosis was measured using the PE-conjugated monoclonalactive caspase-3 antibody apoptosis kit (Becton Dickenson) following thesupplied protocol. Cells were suspended in growth media (0.5×10⁶cells/ml) and treated with the indicated concentrations of each Sykinhibitor or vehicle control for 24, 48, or 72 hours prior to FACSanalysis. The MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide, a tetrazole) assay (company name) was used as a measure of cellviability and growth, following protocols supplied by the manufacturer.Cells were treated with the indicated concentrations of each Sykinhibitor or vehicle control for 72 hours.

SUDHL-4 and SUDHL-6 cells were previously classified as “BCR-type”(Monti, Savage et al. Blood 105(5): 1851-61 (2005); Polo, Juszczynski etal. Proc Natl Acad Sci U S A 104(9): 3207-12 (2007) and sensitive to Sykinhibition by R406 (Chen, Monti et al. 2008). The Toledo and Karpas-422cell lines that lack BCR and BLNK expression, respectively (Gabay,Ben-Bassat et al. Eur J Haematol 63(3): 180-91 (1999); Sprangers,Feldhahn et al. Oncogene 25(36): 5056-62 (2006), having thereforeadapted to survive independent of BCR signals, were insensitive to R406(Chen, Monti et al. 2008). The proliferation of these cell lines whencultured in the presence or absence of various concentrations of eachSyk inhibitor for 72 hours was tested.

Selective inhibition of Syk was sufficient to induce apoptosis in“BCR-type” NHL cell lines. In a separate experiment, the SUDHL-6 andToledo cells were found to be equally sensitive to induction ofapoptosis by 72 h treatment with 1 μM PMA. These data demonstrate thespecific requirement of Syk in the survival of certain NHL cell lines.

Example 161 Xenograft Studies and Tumor and Plasma ConcentrationAnalysis

Syk Inhibition Protects Against Tumor Formation in a Xenograft MouseModel. Mice were received (company) and acclimated in-house at leastthree days prior to use. Ramos cells (3×106) were injectedsubcutaneously into the hind flank area of conscious mice using a 27gauge needle in an injection volume of less than 0.5 ml. Followinginjection, mice were randomized into treatment groups (n=15) and dosedtwice daily by oral gavage with vehicle or 10, 15, or 20 mg/kg of theSyk inhibitor. Body weights were obtained at least once per week andcaliper measurements of tumors were determined twice per week beginningwhen palpable tumors were formed until the end of the study. Tumorvolume was assessed by caliper measurement using a formula [maximumlength×width×height×π/6]. Twice daily dosing of vehicle or the Sykinhibitor continued until the vehicle or any treatment group exhibitedtumors that exceeded 1.5 grams in size. At the time of termination (5weeks post Ramos innoculation) the mice were anesthetized with aketamine cocktail. A blood sample was obtained for CBC and plasmaconcentration determination via cardiac puncture and the mice wereeuthanized via cervical dislocation. Tumors were then be excised andweighed. One half of the tumor was snap frozen in liquid nitrogen fordetermination of concentration of the Syk inhibitor in the tumor tissueand the other half was placed in 10% buffered formalin for histologicalinvestigation.

The effect of Syk inhibition on Ramos tumor formation in a xenograftmouse model was assessed. Mice were dosed twice daily with 10, 15, or 20mg/kg the Syk inhibitor or vehicle control beginning the day of tumorcell inoculation. Caliper measurements were initiated when tumors beganto form, approximately three weeks post-tumor inoculation, and repeatedevery third day until termination of the study. The study was terminatedwhen tumor weights began reaching approximately 1.5 mg, at which timetumors were excised and weighed. Tumor and plasma samples were subjectedto pharmacokinetic analysis.

Each tumor sample was homogenized in 3 ml of saline per gram of tumorusing the Kontes® Microtube Pellet Pestle® Rods and Motor (Kimble Chase,Vineland, N.J.). Plasma and tumor samples were analyzed for the Sykinhibitor concentration using a liquid chromatography tandem massspectrometer (LC/MS/MS). In brief, plasma and tumor samples wereprocessed in a 96-well Captiva™ filter plate (0.2 nm, Varian, Inc., PaloAlto, Calif.). Aliquots of plasma and homogenized tumor samples wereprecipitated with acetonitrile containing 200 ng/mL of:

the internal standard. The mixture was vortexed and refrigerated at 4°C. for 30 minutes to allow complete protein precipitation. The mixturewas filtered into a 96-well collection plate. The filtrate was injectedonto a Sciex API3000 LC/MS/MS equipped with a turbo-ion spray source.The Syk inhibitor and compound A were separated on a Phenomenex Luna 5μHILIC column (4.6×100 mm, 5 mm; Phenomenex, Torrance, Calif.). A mobilephase gradient mixture of 10% mobile phase A (0.1% formic acid in water)and 90% mobile phase B (0.1% formic acid in 90% acetonitrile, 10% water)to 65% mobile phase B was programmed over 1.1 minutes followed by agradient of mobile phase B from 65% to 90% over 0.01 minutes. The peakareas of the m/z 394/360 product ion of the Syk inhibitor were measuredagainst those of the m/z 357/295 product ion of the Syk inhibitor(internal standard) in positive ion mode. The analytical range was 2 to5000 ng/ml.

Pharmacokinetic analysis revealed that at steady-state, tumorconcentrations of the Syk inhibitor followed the concentration-timeprofiles seen with plasma in the 10, 15, and 20 mg/kg dose groups.Nonlinear increases in Cmax, AUC (0-8), and tumor Cmin were observed asthe dose was increased, but a dose-proportional increase in plasma Cminwas noted. Mean Cmax and AUC (0-8) in plasma was at least 2-fold greaterthan that in tumor for all doses examined; however, mean nadirconcentrations (Cmin) were higher in tumor than in plasma, indicatingaccumulation of the Syk inhibitor in the tumor compartment. Tumor/plasmaratios determined from Cmax and AUC (0-8) were similar across thevarious dose groups. Tumor concentrations were sustained above 60, 170,and 640 nM over the entire dosing interval at steady-state for the Sykinhibitor at 10, 15, and 20 mg/kg, respectively.

Mice dosed with all three concentrations of the Syk inhibitor wereprotected from Ramos tumor growth in vivo. This was first evident fromcaliper measurements (data not shown), which revealed a reduced rate oftumor growth in the presence of the Syk inhibitor. Upon studycompletion, mice were euthanized and tumors excised and weighed.Consistent with caliper measurements, a statistically significantreduction in average tumor weight was achieved in all dosing groups,relative to vehicle control. These data reveal that sub-micromolarconcentrations of the Syk inhibitor can prevent tumor formation by anaggressive NHL cell line in mice.

Mice dosed with the Syk inhibitor did not present with reduced numbersin any subset of white blood cells. In fact, the only effect observedwas an increase in the number of lymphocytes in mice treated with 15mg/kg the Syk inhibitor which was not repeated in mice dosed with 10 or20 mg/kg. The relative percent of each cell subtype analyzed was alsounaffected by the Syk inhibitor (data not shown). On average, micetreated with vehicle control had a 9.45% increase in body weight. Micetreated with 10, 15, and 20 mg/kg the Syk inhibitor, on the other hand,had on average 0.27% increase, 1.67% decrease, and 2.27% decrease inbody weight, respectively, over the course of the study. There was norelationship, however, between % change in body weight and tumor growth(R2=0.27). These data suggest that the inhibition of tumor growth wasindeed mediated by suppression of Syk activity.

The Syk-specific inhibitor the Syk inhibitor was also tested foractivity in a Ramos tumor mouse xenograft model. At all theconcentrations tested, statistically significant reductions in tumorgrowth were observed in mice dosed BID with the Syk inhibitor. Thelowest concentration tested was 10 mg/kg, achieving tumor concentrationsranging from 64 to 140 nM over the course of the day. Suppression oftumor growth at these concentrations in vivo is consistent withconcentrations of <125 nM found to suppress BCR-induced Ca2+ flux anddistal BCR signaling to pERK Y204. The selective pharmacologicalinhibition of Syk results in effects on the proliferations and survivalof NHL cell lines. These data suggest that the selective targeting ofSyk may similarly have clinical benefit in a variety of B-cellproliferative disorders.

As detailed herein, Syk has been implicated experimentally in B celldevelopment, proliferation, and survival. Moreover, Syk is implicated asan oncogene. Expression of constitutively active Syk in adoptivelytransferred bone marrow cells induces leukemia in mice, andover-activity of Syk is associated with a variety of lymphomas in humansGiven the role of Syk in B cell biology, its selective inhibition may besufficient to provide clinical benefit in B cell proliferativedisorders, while reducing toxicities that may arise due to suppressionof other off-target kinases.

The present invention provides a number of embodiments. It is apparentthat the examples may be altered to provide other embodiments of thisinvention. Therefore, it will be appreciated that the scope of thisinvention is to be defined by the appended claims rather than by thespecific embodiments, which have been represented by way of example.

All of the above U.S. patents, U.S. patent application publications,U.S. patent applications, foreign patents, foreign patent applicationsand non-patent publications referred to in this specification and/orlisted in the Application Data Sheet, are incorporated herein byreference, in their entirety. From the foregoing it will be appreciatedthat, although specific embodiments of the invention have been describedherein for purposes of illustration, various modifications may be madewithout deviating from the spirit and scope of the invention.Accordingly, the invention is not limited except as by the appendedclaims.

1.-59. (canceled)
 60. A compound having the formula II:

or a tautomer or a pharmaceutically acceptable salt thereof, wherein:R^(1c) is selected from the group consisting of (a) C₃₋₈cycloalkyl, and(b) aryl, each of which is optionally substituted with from 1 to 2substituents selected from the group consisting of C₁₋₈alkyl, amino andhydroxyl; or is taken together with R^(2c) to form a heterocyclic ring,containing 1-3 heteroatoms, including N, O, S, optionally substitutedwith from 1 to 2 substituents, each of which is independently selectedfrom the group consisting of aminocarbonylC₁₋₈alkylene,C₁₋₈alkoxycarbonyl, aminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene,C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene, amino,aminocarbonylaminocarbonylamino, arylC₁₋₈alkoxycarbonylamino,aminocarbonylamino and oxo; R^(2c) is H or is taken together with R^(1c)to form a heterocyclic ring, optionally cyanoC₁₋₈alkylcarbonyl,cyanoC₁₋₆alkylcarbonylamino substituted with from 1 to 2 substituentsindependently selected from the group consisting ofaminocarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonyl, aminoC₁₋₈alkylene,hydroxyC₁₋₈alkylene, C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene, amino,aminocarbonylaminocarbonylamino, arylC₁₋₈alkoxycarbonylamino,aminocarbonylamino and oxo; R^(3c) is H or taken together with R^(4d) toform a heterocyclic or heteroaryl ring, each of which is optionallysubstituted with from 1 to 2 substituents independently selected fromthe group consisting of C₁₋₈alkyl, C₁₋₈alkylheterocyclyl,aminoC₁₋₈alkylene, aminoaryl, hydroxyC₁₋₈alkylene, aminocarbonyl,C₁₋₈alkoxycarbonyl, amino, imino, C₁₋₈alkylcarbonylamino, oxo, halo,aryl, heterocyclyl, heterocyclylC₁₋₈alkylene andC₁₋₈alkylheterocyclylC₁₋₈alkylene; R^(4d) is independently selected fromthe group consisting of (a) C₃₋₈cycloalkyl, (b) aryl, (c) heteroaryl,and (d) heterocyclyl; each of which is optionally substituted with from1 to 3 substituents R^(4e), R^(4f) and —Z^(1c)R^(4g), each of which isindependently selected from the group consisting of C₁₋₈alkyl,aminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene, aminocarbonyl,C₁₋₈alkoxycarbonyl, carboxy, amino, imino, C₁₋₈alkylcarbonylamino, oxo,halo, aryl, heterocyclyl and heterocyclylC₁₋₈alkylene, provided that atleast one of R^(1c) and R^(2c) or R^(3c) and R^(4d) are combined withthe nitrogen to which each is attached to form a heterocyclyl orheteroaryl ring; and R^(7c) is selected from the group consisting of H,C₁₋₈alkyl, C₂₋₈alkynyl, cyano, aminocarbonyl, nitro, C₁₋₈alkoxy,halogen, aryl, heteroaryl and C₃₋₈cycloalkyl.
 61. The compound of claim60 having the formula IIa:

or a tautomer or a pharmaceutically acceptable salt thereof.
 62. Thecompound of any one of claims 60 to 61, wherein R^(3c) is H.
 63. Thecompound of any one of claims 60 to 61, wherein R^(4d) is phenyl. 64.The compound of any one of claims 60 to 61, wherein R^(4d) is indazyl.65. The compound of any one of claims 60 to 61, wherein R^(4d) iscyclohexyl.
 66. The compound of any one of claims 60 to 61, whereinR^(4d) is dihydroindoyl.
 67. The compound of any one of claims 60 to 61,wherein R^(3c) is taken together with R^(4d) and the nitrogen to whichis each is attached to form heterocyclic ring selected from the groupconsisting of pyridinyl, imidazolyl, tetrahydroimidazoyl, indazolyl,piperidinyl, piperazinyl, pyrrolidinyl, triazoyl and benzamidazoyl. 68.A compound of claim 61 or a pharmaceutically acceptable salt thereof,wherein the moiety R^(4d)R^(3c)N— is selected from the group consistingof:

the dashed lined indicates a single or double bond; and the wavy lineindicates the point of attachment to the remainder of the molecule. 69.The compound of claim 60 having the formula IIb:

or a tautomer or a pharmaceutically acceptable salt thereof, wherein:R^(7c) is halo; R^(1c) and R^(2c) are taken together to form aheterocyclic ring, optionally substituted with from 1 to 2 substituentsindependently selected from the group consisting of:aminocarbonylC₁₋₈alkylene, C₁₋₈alkoxycarbonyl, aminoC₁₋₈alkylene,hydroxyC₁₋₈alkylene, C₁₋₈alkoxycarbonylaminoC₁₋₈alkylene, amino,aminocarbonylaminocarbonylamino, arylC₁₋₈alkoxycarbonylamino,aminocarbonylamino and oxo; Z^(1c) is heterocyclyl, —N(C₁₋₄alkyl)-,—SO₂— or —CO—; R^(4g) is selected from the group consisting of H,C₁₋₈alkoxy, C₁₋₈alkylcarbonyl, C₁₋₈alkylcarbonylamino, amino,aminocarbonylaminoC₁₋₈alkylene, aminocarbonyl and aminosulfonyl; eachR^(4e) and R^(4f) is independently selected from the group consisting ofH—, C₁₋₈alkoxy and C₁₋₈alkylcarbonyl; or can be taken together with—Z^(1c)—R^(4g) and the benzene ring to which each is attached to form afused heterocyclic ring system, optionally substituted with from 1 to 2oxo substituents, halogens, C₁₋₈alkyl, C₁₋₈alkyl.
 70. The compound ofclaim 69, wherein the moiety

is selected from the group consisting of:

71.-79. (canceled)
 80. The compound of claim 60 having the formulaselected from the group consisting of:

wherein Y^(2b) is N, NH, C or CH; R^(2d) is H,C₁₋₈alkylaminoC₁₋₈alkylene, aminocarbonyl, aminocarbonylamino,aminocarbonylaminocarbonylamino, amino, oxo, aminoC₁₋₈alkylene,aminocarbonylaminoC₁₋₈alkylene, hydroxyC₁₋₈alkylene,arylalkoxycarbonylamino; cyanoC₁₋₈alkylenecarbonyl,cyanoC₁₋₈alkylenecarbonylamino, hydroxyl, C₁₋₈alkoxycarbonyl,alkoxycarbonylC₁₋₈alkylene, optionally substituted by amino; and R^(7C)is H, halo, aminocarbonyl, cycloalkyl, cyano or pyridinyl; R⁹ is H orC₁₋₈alkyl; R¹⁰ is H, C₁₋₈alkyl, C₁₋₈alkoxy,C₁₋₈alkoxycarbonylC₁₋₈alkylene, aminoC₁₋₈alkylene,aminocarbonylC₁₋₈alkylene, carboxyC₁₋₈alkylene, C₃₋₈cycloalkyl andhydroxyC₁₋₈alkylene; m is 0 or 1; and q is 0 or
 1. 81. The compound ofclaim 60, wherein R^(7c) is selected from the group consisting of F, Cl,Br, cyano and aminocarbonyl.
 82. The compound of claim 60, whereinR^(7c) is CONH₂ or F.
 83. The compound of claim 60 having the formula:

or a tautomer or a pharmaceutically acceptable salt thereof. 84.-88.(canceled)
 89. A composition comprising a compound of any one of thepreceding claims in combination with a pharmaceutically acceptablecarrier or diluent.
 90. A method for inhibiting syk, JAK kinase or asignal transduction pathway mediated at least in part by syk or JAKkinase activity comprising the step of contacting a cell with a compoundof any one of the preceding claims.
 91. A method for treating acondition or disorder mediated at least in part by syk or JAK kinaseactivity in a subject comprising the step of administering to a subjectin need of such treatment a therapeutically effective amount of acomposition of claim
 89. 92. The method of claim 89, wherein thecondition or disorder is selected from the group consisting ofcardiovascular disease, inflammatory disease, immune-related disease andcell proliferative disorder. 93.-102. (canceled)
 103. A kit comprising acomposition of claim 89, packaging and instructions for use.